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  • 1. Acero Sanchez, Josep Ll.
    et al.
    Joda, Hamdi
    Henry, Olivier Y. F.
    Solnestam, Beata W.
    Kvastad, Linda
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Sahlén, Pelin
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lundeberg, Joakim
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Laddach, Nadja
    Ramakrishnan, Dheeraj
    Riley, Ian
    Schwind, Carmen
    Latta, Daniel
    O'Sullivan, Ciara K.
    Electrochemical Genetic Profiling of Single Cancer Cells2017In: Analytical Chemistry, ISSN 0003-2700, E-ISSN 1520-6882, Vol. 89, no 6, p. 3378-3385Article in journal (Refereed)
    Abstract [en]

    Recent understandings in the development and spread of cancer have led to the realization of novel single cell analysis platforms focused on circulating tumor cells (CTCs). A simple, rapid, and inexpensive analytical platform capable of providing genetic information on these rare cells is highly desirable to support clinicians and researchers alike to either support the selection or adjustment of therapy or provide fundamental insights into cell function and cancer progression mechanisms. We report on the genetic profiling of single cancer cells, exploiting a combination of multiplex ligation-dependent probe amplification (MLPA) and electrochemical detection. Cells were isolated using laser capture and lysed, and the mRNA was extracted and transcribed into DNA. Seven markers were amplified by MLPA, which allows for the simultaneous amplification of multiple targets with a single primer pair, using MLPA probes containing unique barcode sequences. Capture probes complementary to each of these barcode sequences were immobilized on a printed circuit board (PCB) manufactured electrode array and exposed to single-stranded MLPA products and subsequently to a single stranded DNA reporter probe bearing a HRP molecule, followed by substrate addition and fast electrochemical pulse amperometric detection. We present asimple, rapid, flexible, and inexpensive approach for the simultaneous quantification of multiple breast cancer related mRNA markers, with single tumor cell sensitivity.

  • 2.
    Akan, Pelin
    et al.
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Alexeyenko, Andrey
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Costea, Paul Igor
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Hedberg, Lilia
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Werne Solnestam, Beata
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lundin, Sverker
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Hallman, Jimmie
    Lundberg, Emma
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Uhlén, Mathias
    KTH, School of Biotechnology (BIO), Proteomics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lundeberg, Joakim
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Comprehensive analysis of the genome transcriptome and proteome landscapes of three tumor cell lines2012In: Genome Medicine, ISSN 1756-994X, E-ISSN 1756-994X, Vol. 4, p. 86-Article in journal (Refereed)
    Abstract [en]

    We here present a comparative genome, transcriptome and functional network analysis of three human cancer cell lines (A431, U251MG and U2OS), and investigate their relation to protein expression. Gene copy numbers significantly influenced corresponding transcript levels; their effect on protein levels was less pronounced. We focused on genes with altered mRNA and/or protein levels to identify those active in tumor maintenance. We provide comprehensive information for the three genomes and demonstrate the advantage of integrative analysis for identifying tumor-related genes amidst numerous background mutations by relating genomic variation to expression/protein abundance data and use gene networks to reveal implicated pathways.

  • 3.
    Akan, Pelin
    et al.
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Biotechnology (BIO).
    Stranneheim, Henrik
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Biotechnology (BIO).
    Lexow, Preben
    LingVitae, Oslo.
    Lundeberg, Joakim
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Biotechnology (BIO).
    Design and assessment of binary DNA for nanopore sequencing2010In: Genome biology, ISSN 1474-760X, Vol. 11, p. P4-Article in journal (Other academic)
  • 4.
    Anil, Anandashankar
    et al.
    KTH, School of Biotechnology (BIO). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Spalinskas, Rapolas
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Biotechnology (BIO).
    Åkerborg, Örjan
    KTH, School of Biotechnology (BIO). KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Sahlén, Pelin
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Biotechnology (BIO).
    HiCapTools: a software suite for probe design and proximity detection for targeted chromosome conformation capture applications2018In: Bioinformatics, ISSN 1367-4803, E-ISSN 1367-4811, Vol. 34, no 4, p. 675-677Article in journal (Refereed)
    Abstract [en]

    Folding of eukaryotic genomes within nuclear space enables physical and functional contacts between regions that are otherwise kilobases away in sequence space. Targeted chromosome conformation capture methods (T2C, chi-C and HiCap) are capable of informing genomic contacts for a subset of regions targeted by probes. We here present HiCapTools, a software package that can design sequence capture probes for targeted chromosome capture applications and analyse sequencing output to detect proximities involving targeted fragments. Two probes are designed for each feature while avoiding repeat elements and non-unique regions. The data analysis suite processes alignment files to report genomic proximities for each feature at restriction fragment level and is isoform-aware for gene features. Statistical significance of contact frequencies is evaluated using an empirically derived background distribution. Targeted chromosome conformation capture applications are invaluable for locating target genes of disease-associated variants found by genome-wide association studies. Hence, we believe our software suite will prove to be useful for a wider user base within clinical and functional applications.

  • 5.
    Banfi, Cristina
    et al.
    Ctr Cardiol Monzino IRCCS, Unit Funct Prote Metabol & Network Anal, I-20138 Milan, Italy..
    Mallia, Alice
    Ctr Cardiol Monzino IRCCS, Unit Funct Prote Metabol & Network Anal, I-20138 Milan, Italy.;Univ Pavia, Dipartimento Biol & Biotecnol Lazzaro Spallanzani, I-27100 Pavia, Italy..
    Ghilardi, Stefania
    Ctr Cardiol Monzino IRCCS, Unit Funct Prote Metabol & Network Anal, I-20138 Milan, Italy..
    Brioschi, Maura
    Ctr Cardiol Monzino IRCCS, Unit Funct Prote Metabol & Network Anal, I-20138 Milan, Italy..
    Gianazza, Erica
    Ctr Cardiol Monzino IRCCS, Unit Funct Prote Metabol & Network Anal, I-20138 Milan, Italy..
    Eligini, Sonia
    Ctr Cardiol Monzino IRCCS, Unit Funct Prote Metabol & Network Anal, I-20138 Milan, Italy..
    Sahlén, Pelin
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Baetta, Roberta
    Ctr Cardiol Monzino IRCCS, Unit Funct Prote Metabol & Network Anal, I-20138 Milan, Italy..
    Prenylcysteine Oxidase 1 Is a Key Regulator of Adipogenesis2023In: Antioxidants, ISSN 2076-3921, Vol. 12, no 3, p. 542-, article id 542Article in journal (Refereed)
    Abstract [en]

    The process of adipogenesis involves the differentiation of preadipocytes into mature adipocytes. Excessive adipogenesis promotes obesity, a condition that increasingly threatens global health and contributes to the rapid rise of obesity-related diseases. We have recently shown that prenylcysteine oxidase 1 (PCYOX1) is a regulator of atherosclerosis-disease mechanisms, which acts through mechanisms not exclusively related to its pro-oxidant activity. To address the role of PCYOX1 in the adipogenic process, we extended our previous observations confirming that Pcyox1(-/-)/Apoe(-/-) mice fed a high-fat diet for 8 or 12 weeks showed significantly lower body weight, when compared to Pcyox1(+/+)/Apoe(-/-) mice, due to an evident reduction in visceral adipose content. We herein assessed the role of PCYOX1 in adipogenesis. Here, we found that PCYOX1 is expressed in adipose tissue, and, independently from its pro-oxidant enzymatic activity, is critical for adipogenesis. Pcyox1 gene silencing completely prevented the differentiation of 3T3-L1 preadipocytes, by acting as an upstream regulator of several key players, such as FABP4, PPAR gamma, C/EBP alpha. Proteomic analysis, performed by quantitative label-free mass spectrometry, further strengthened the role of PCYOX1 in adipogenesis by expanding the list of its downstream targets. Finally, the absence of Pcyox1 reduces the inflammatory markers in adipose tissue. These findings render PCYOX1 a novel adipogenic factor with possible pathophysiological or therapeutic potential.

  • 6.
    Bjorn, Niclas
    et al.
    Linköping Univ, Div Drug Res, Dept Biomed & Clin Sci, Clin Pharmacol, Linköping, Sweden..
    Badam, Tejaswi Venkata Satya
    Linköping Univ, Dept Phys Chem & Biol, Bioinformat, Linköping, Sweden.;Univ Skövde, Sch Biosci, Syst Biol Res Ctr, Skövde, Sweden..
    Spalinskas, Rapolas
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Branden, Eva
    Gävle Cent Hosp, Dept Resp Med, Gävle, Sweden.;Uppsala Univ Reg Gävleborg, Ctr Res & Dev, Gävle, Sweden..
    Koyi, Hirsh
    Gävle Cent Hosp, Dept Resp Med, Gävle, Sweden.;Uppsala Univ Reg Gävleborg, Ctr Res & Dev, Gävle, Sweden..
    Lewensohn, Rolf
    Karolinska Univ Hosp, Thorac Oncol Unit, Tema Canc, Stockholm, Sweden.;Karolinska Inst, Dept Oncol Pathol, Stockholm, Sweden..
    De Petris, Luigi
    Karolinska Univ Hosp, Thorac Oncol Unit, Tema Canc, Stockholm, Sweden.;Karolinska Inst, Dept Oncol Pathol, Stockholm, Sweden..
    Lubovac-Pilav, Zelmina
    Univ Skövde, Sch Biosci, Syst Biol Res Ctr, Skövde, Sweden..
    Sahlén, Pelin
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Lundeberg, Joakim
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Gustafsson, Mika
    Linköping Univ, Dept Phys Chem & Biol, Bioinformat, Linköping, Sweden..
    Gréen, Henrik
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab. Linköping Univ, Div Drug Res, Dept Biomed & Clin Sci, Clin Pharmacol, Linköping, Sweden.; Natl Board Forens Med, Dept Forens Genet & Forens Toxicol, Linköping, Sweden..
    Whole-genome sequencing and gene network modules predict gemcitabine/carboplatin-induced myelosuppression in non-small cell lung cancer patients2020In: NPJ SYSTEMS BIOLOGY AND APPLICATIONS, ISSN 2056-7189, Vol. 6, no 1, article id 25Article in journal (Refereed)
    Abstract [en]

    Gemcitabine/carboplatin chemotherapy commonly induces myelosuppression, including neutropenia, leukopenia, and thrombocytopenia. Predicting patients at risk of these adverse drug reactions (ADRs) and adjusting treatments accordingly is a long-term goal of personalized medicine. This study used whole-genome sequencing (WGS) of blood samples from 96 gemcitabine/carboplatin-treated non-small cell lung cancer (NSCLC) patients and gene network modules for predicting myelosuppression. Association of genetic variants in PLINK found 4594, 5019, and 5066 autosomal SNVs/INDELs withp <= 1 x 10(-3)for neutropenia, leukopenia, and thrombocytopenia, respectively. Based on the SNVs/INDELs we identified the toxicity module, consisting of 215 unique overlapping genes inferred from MCODE-generated gene network modules of 350, 345, and 313 genes, respectively. These module genes showed enrichment for differentially expressed genes in rat bone marrow, human bone marrow, and human cell lines exposed to carboplatin and gemcitabine (p < 0.05). Then using 80% of the patients as training data, random LASSO reduced the number of SNVs/INDELs in the toxicity module into a feasible prediction model consisting of 62 SNVs/INDELs that accurately predict both the training and the test (remaining 20%) data with high (CTCAE 3-4) and low (CTCAE 0-1) maximal myelosuppressive toxicity completely, with the receiver-operating characteristic (ROC) area under the curve (AUC) of 100%. The present study shows how WGS, gene network modules, and random LASSO can be used to develop a feasible and tested model for predicting myelosuppressive toxicity. Although the proposed model predicts myelosuppression in this study, further evaluation in other studies is required to determine its reproducibility, usability, and clinical effect.

  • 7.
    Björn, Niclas
    et al.
    Linköping University, Department of Medical and Health Sciences, Division of Drug Research. Linköping University, Faculty of Medicine and Health Sciences..
    Pradhananga, Sailendra
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Sigurgeirsson, Benjamin
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lundeberg, Joakim
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Gréen, Henrik
    Linköping University, Department of Medical and Health Sciences, Division of Drug Research. Linköping University, Faculty of Medicine and Health Sciences..
    Sahlén, Pelin
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Comparison of Variant Calls from Whole Genome and Whole Exome Sequencing Data Using Matched Samples2018In: Journal of Next Generation Sequencing & Applications, ISSN 2469-9853, Vol. 5, no 1, p. 1-8Article in journal (Refereed)
    Abstract [en]

    Whole exome sequencing (WES) has been extensively used in genomic research. As sequencing costs decline it is being replaced by whole genome sequencing (WGS) in large-scale genomic studies, but more comparative information on WES and WGS datasets would be valuable. Thus, we have extensively compared variant calls obtained from WGS and WES of matched germline DNA samples from 96 lung cancer patients. WGS provided more homogeneous coverage with higher genotyping quality, and identified more variants, than WES, regardless of exome coverage depth. It also called more reference variants, reflecting its power to call rare variants, and more heterozygous variants that met applied quality criteria, indicating that WGS is less prone to allelic drop outs. However, increasing WES coverage reduced the discrepancy between the WES and WGS results. We believe that as sequencing costs further decline WGS will become the method of choice even for research confined to the exome.

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  • 8.
    Cavalli, M.
    et al.
    Uppsala Univ, Uppsala, Sweden..
    Baltzer, N.
    Uppsala Univ, Uppsala, Sweden..
    Umer, H. M.
    Uppsala Univ, Uppsala, Sweden..
    Grau, J.
    Martin Luther Univ HalleWittenberg, Halle, Germany..
    Lemnian, I.
    Martin Luther Univ HalleWittenberg, Halle, Germany..
    Pan, G.
    Uppsala Univ, Uppsala, Sweden..
    Wallerman, O.
    Uppsala Univ, Uppsala, Sweden..
    Spalinskas, Rapolas
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Sahlén, Pelin
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Grosse, I.
    Martin Luther Univ HalleWittenberg, Halle, Sweden..
    Komorowski, J.
    Uppsala Univ, Uppsala, Sweden..
    Wadelius, C.
    Uppsala Univ, Uppsala, Sweden..
    Allele specific chromatin signals, 3D interactions, and refined motif predictions for immune and B cell related diseases2019In: European Journal of Human Genetics, ISSN 1018-4813, E-ISSN 1476-5438, Vol. 27, p. 611-611Article in journal (Other academic)
  • 9. Cavalli, M.
    et al.
    Diamanti, K.
    Pan, G.
    Spalinskas, Rapolas
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Kumar, C.
    Deshmukh, A. S.
    Mann, M.
    Sahlén, Pelin
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Komorowski, J.
    Wadelius, C.
    A Multi-Omics Approach to Liver Diseases: Integration of Single Nuclei Transcriptomics with Proteomics and HiCap Bulk Data in Human Liver2020In: Omics, ISSN 1536-2310, E-ISSN 1557-8100, Vol. 24, no 4, p. 180-194Article in journal (Refereed)
    Abstract [en]

    The liver is the largest solid organ and a primary metabolic hub. In recent years, intact cell nuclei were used to perform single-nuclei RNA-seq (snRNA-seq) for tissues difficult to dissociate and for flash-frozen archived tissue samples to discover unknown and rare cell subpopulations. In this study, we performed snRNA-seq of a liver sample to identify subpopulations of cells based on nuclear transcriptomics. In 4282 single nuclei, we detected, on average, 1377 active genes and we identified seven major cell types. We integrated data from 94,286 distal interactions (p < 0.05) for 7682 promoters from a targeted chromosome conformation capture technique (HiCap) and mass spectrometry proteomics for the same liver sample. We observed a reasonable correlation between proteomics and in silico bulk snRNA-seq (r = 0.47) using tissue-independent gene-specific protein abundancy estimation factors. We specifically looked at genes of medical importance. The DPYD gene is involved in the pharmacogenetics of fluoropyrimidine toxicity and some of its variants are analyzed for clinical purposes. We identified a new putative polymorphic regulatory element, which may contribute to variation in toxicity. Hepatocellular carcinoma (HCC) is the most common type of primary liver cancer and we investigated all known risk genes. We identified a complex regulatory landscape for the SLC2A2 gene with 16 candidate enhancers. Three of them harbor somatic motif breaking and other mutations in HCC in the Pan Cancer Analysis of Whole Genomes dataset and are candidates to contribute to malignancy. Our results highlight the potential of a multi-omics approach in the study of human diseases.

  • 10.
    Cavalli, Marco
    et al.
    Uppsala Univ, Dept Immunol Genet & Pathol, Sci Life Lab, Uppsala, Sweden..
    Baltzer, Nicholas
    Uppsala Univ, Dept Cell & Mol Biol Computat Biol & Bioinformat, Uppsala, Sweden..
    Umer, Husen M.
    Uppsala Univ, Dept Cell & Mol Biol Computat Biol & Bioinformat, Uppsala, Sweden..
    Grau, Jan
    Martin Luther Univ Halle Wittenberg, Inst Comp Sci, Halle, Germany..
    Lemnian, Ioana
    Martin Luther Univ Halle Wittenberg, Inst Comp Sci, Halle, Germany..
    Pan, Gang
    Uppsala Univ, Dept Immunol Genet & Pathol, Sci Life Lab, Uppsala, Sweden..
    Wallerman, Ola
    Uppsala Univ, Dept Immunol Genet & Pathol, Sci Life Lab, Uppsala, Sweden..
    Spalinskas, Rapolas
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Sahlén, Pelin
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Grosse, Ivo
    Martin Luther Univ Halle Wittenberg, Inst Comp Sci, Halle, Germany.;German Ctr Integrat Biodivers Res iDiv, Leipzig, Germany..
    Komorowski, Jan
    Uppsala Univ, Dept Cell & Mol Biol Computat Biol & Bioinformat, Uppsala, Sweden.;Polish Acad Sci, Inst Comp Sci, Warsaw, Poland..
    Wadelius, Claes
    Uppsala Univ, Dept Immunol Genet & Pathol, Sci Life Lab, Uppsala, Sweden..
    Allele specific chromatin signals, 3D interactions, and motif predictions for immune and B cell related diseases2019In: Scientific Reports, E-ISSN 2045-2322, Vol. 9, article id 2695Article in journal (Refereed)
    Abstract [en]

    Several Genome Wide Association Studies (GWAS) have reported variants associated to immune diseases. However, the identified variants are rarely the drivers of the associations and the molecular mechanisms behind the genetic contributions remain poorly understood. ChIP-seq data for TFs and histone modifications provide snapshots of protein-DNA interactions allowing the identification of heterozygous SNPs showing significant allele specific signals (AS-SNPs). AS-SNPs can change a TF binding site resulting in altered gene regulation and are primary candidates to explain associations observed in GWAS and expression studies. We identified 17,293 unique AS-SNPs across 7 lymphoblastoid cell lines. In this set of cell lines we interrogated 85% of common genetic variants in the population for potential regulatory effect and we identified 237 AS-SNPs associated to immune GWAS traits and 714 to gene expression in B cells. To elucidate possible regulatory mechanisms we integrated long-range 3D interactions data to identify putative target genes and motif predictions to identify TFs whose binding may be affected by AS-SNPs yielding a collection of 173 AS-SNPs associated to gene expression and 60 to B cell related traits. We present a systems strategy to find functional gene regulatory variants, the TFs that bind differentially between alleles and novel strategies to detect the regulated genes.

  • 11.
    Costea, Paul Igor
    et al.
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lundeberg, Joakim
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Akan, Pelin
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    TagGD: Fast and Accurate Software for DNA Tag Generation and Demultiplexing2013In: PLOS ONE, E-ISSN 1932-6203, Vol. 8, no 3, p. e57521-Article in journal (Refereed)
    Abstract [en]

    Multiplexing is of vital importance for utilizing the full potential of next generation sequencing technologies. We here report TagGD (DNA-based Tag Generator and Demultiplexor), a fully-customisable, fast and accurate software package that can generate thousands of barcodes satisfying user-defined constraints and can guarantee full demultiplexing accuracy. The barcodes are designed to minimise their interference with the experiment. Insertion, deletion and substitution events are considered when designing and demultiplexing barcodes. 20,000 barcodes of length 18 were designed in 5 minutes and 2 million barcoded Illumina HiSeq-like reads generated with an error rate of 2% were demultiplexed with full accuracy in 5 minutes. We believe that our software meets a central demand in the current high-throughput biology and can be utilised in any field with ample sample abundance. The software is available on GitHub (https://github.com/pelinakan/UBD.git).

  • 12.
    Freiholtz, David
    et al.
    Section of Cardiothoracic Surgery, Department of Molecular Medicine and Surgery, Karolinska Institutet, Stockholm, Sweden; Division of Cardiovascular Medicine, Center for Molecular Medicine, Department of Medicine, Karolinska Institutet, Stockholm, Karolinska University Hospital, Solna, Sweden.
    Bergman, Otto
    Division of Cardiovascular Medicine, Center for Molecular Medicine, Department of Medicine, Karolinska Institutet, Stockholm, Karolinska University Hospital, Solna, Sweden.
    Pradhananga, Sailendra
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lång, Karin
    Division of Cardiovascular Medicine, Center for Molecular Medicine, Department of Medicine, Karolinska Institutet, Stockholm, Karolinska University Hospital, Solna, Sweden.
    Poujade, Flore Anne
    Division of Cardiovascular Medicine, Center for Molecular Medicine, Department of Medicine, Karolinska Institutet, Stockholm, Karolinska University Hospital, Solna, Sweden.
    Granath, Carl
    Section of Cardiothoracic Surgery, Department of Molecular Medicine and Surgery, Karolinska Institutet, Stockholm, Sweden; Division of Cardiovascular Medicine, Center for Molecular Medicine, Department of Medicine, Karolinska Institutet, Stockholm, Karolinska University Hospital, Solna, Sweden.
    Olsson, Christian
    Section of Cardiothoracic Surgery, Department of Molecular Medicine and Surgery, Karolinska Institutet, Stockholm, Sweden.
    Franco-Cereceda, Anders
    Section of Cardiothoracic Surgery, Department of Molecular Medicine and Surgery, Karolinska Institutet, Stockholm, Sweden.
    Sahlén, Pelin
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Eriksson, Per
    Division of Cardiovascular Medicine, Center for Molecular Medicine, Department of Medicine, Karolinska Institutet, Stockholm, Karolinska University Hospital, Solna, Sweden.
    Björck, Hanna M.
    Division of Cardiovascular Medicine, Center for Molecular Medicine, Department of Medicine, Karolinska Institutet, Stockholm, Karolinska University Hospital, Solna, Sweden.
    SPP1/osteopontin: a driver of fibrosis and inflammation in degenerative ascending aortic aneurysm?2023In: Journal of Molecular Medicine, ISSN 0946-2716, E-ISSN 1432-1440, Vol. 101, no 10, p. 1323-1333Article in journal (Refereed)
    Abstract [en]

    Abstract: Degenerative ascending aortic aneurysm (AscAA) is a silent and potentially fatal disease characterized by excessive vascular inflammation and fibrosis. We aimed to characterize the cellular and molecular signature for the fibrotic type of endothelial mesenchymal transition (EndMT) that has previously been described in degenerative AscAA. Patients undergoing elective open-heart surgery for AscAA and/or aortic valve repair were recruited. Gene expression in the intima-media of the ascending aorta was measured in 22 patients with non-dilated and 24 with dilated aortas, and candidate genes were identified. Protein expression was assessed using immunohistochemistry. Interacting distal gene enhancer regions were identified using targeted chromosome conformation capture (HiCap) in untreated and LPS-treated THP1 cells, and the associated transcription factors were analyzed. Differential expression analysis identified SPP1 (osteopontin) as a key gene in the signature of fibrotic EndMT in patients with degenerative AscAA. The aortic intima-media expression of SPP1 correlated with the expression of inflammatory markers, the level of macrophage infiltration, and the aortic diameter. HiCap analysis, followed by transcription factor binding analysis, identified ETS1 as a potential regulator of SPP1 expression under inflammatory conditions. In conclusion, the present findings suggest that SPP1 may be involved in the development of the degenerative type of AscAA. Key messages: In the original manuscript titled “SPP1/osteopontin, a driver of fibrosis and inflammation in degenerative ascending aortic aneurysm?” by David Freiholtz, Otto Bergman, Saliendra Pradhananga, Karin Lång, Flore-Anne Poujade, Carl Granath, Christian Olsson, Anders Franco-Cereceda, Pelin Sahlén, Per Eriksson, and Hanna M Björck, we present novel findings on regulatory factors on osteopontin (SPP1) expression in immune cells involved in degenerative ascending aortic aneurysms (AscAA). The central findings convey: SPP1 is a potential driver of the fibrotic endothelial-to-mesenchymal transition in AscAA.SPP1/osteopontin expression in AscAA is predominately by immune cells.ETS1 is a regulatory transcription factor of SPP1 expression in AscAA immune cells.

  • 13. Hu, M.
    et al.
    Ayub, Q.
    Guerra-Assunção, J. A.
    Long, Q.
    Ning, Z.
    Huang, N.
    Romero, I. G.
    Mamanova, L.
    Akan, Pelin
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Liu, X.
    Coffey, A. J.
    Turner, D. J.
    Swerdlow, H.
    Burton, J.
    Quail, M. A.
    Conrad, D. F.
    Enright, A. J.
    Tyler-Smith, C.
    Xue, Y.
    Exploration of signals of positive selection derived from genotype-based human genome scans using re-sequencing data2012In: Human Genetics, ISSN 0340-6717, E-ISSN 1432-1203, Vol. 131, no 5, p. 665-674Article in journal (Refereed)
    Abstract [en]

    We have investigated whether regions of the genome showing signs of positive selection in scans based on haplotype structure also show evidence of positive selection when sequence-based tests are applied, whether the target of selection can be localized more precisely, and whether such extra evidence can lead to increased biological insights. We used two tools: simulations under neutrality or selection, and experimental investigation of two regions identified by the HapMap2 project as putatively selected in human populations. Simulations suggested that neutral and selected regions should be readily distinguished and that it should be possible to localize the selected variant to within 40 kb at least half of the time. Re-sequencing of two ∼300 kb regions (chr4:158Mb and chr10:22Mb) lacking known targets of selection in HapMap CHB individuals provided strong evidence for positive selection within each and suggested the micro-RNA gene hsa-miR-548c as the best candidate target in one region, and changes in regulation of the sperm protein gene SPAG6 in the other.

  • 14.
    Kvastad, Linda
    et al.
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Werne Solnestam, Beata
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Johansson, Elin
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Nygren, A. O.
    Laddach, N.
    Sahlén, Pelin
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Vickovic, Sanja
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Bendigtsen, S. C.
    Aaserud, M.
    Floer, L.
    Borgen, E.
    Schwind, C.
    Himmelreich, R.
    Latta, D.
    Lundeberg, Joakim
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Single cell analysis of cancer cells using an improved RT-MLPA method has potential for cancer diagnosis and monitoring2015In: Scientific Reports, E-ISSN 2045-2322, Vol. 5, article id 16519Article in journal (Refereed)
    Abstract [en]

    Single cell analysis techniques have great potential in the cancer genomics feld. The detection and characterization of circulating tumour cells are important for identifying metastatic disease at an early stage and monitoring it. This protocol is based on transcript profiling using Reverse Transcriptase Multiplex Ligation-dependent Probe Amplification (RT-MLPA), which is a specific method for simultaneous detection of multiple mRNA transcripts. Because of the small amount of (circulating) tumour cells, a pre-amplification reaction is performed after reverse transcription to generate a sufficient number of target molecules for the MLPA reaction. We designed a highly sensitive method for detecting and quantifying a panel of seven genes whose expression patterns are associated with breast cancer, and optimized the method for single cell analysis. For detection we used a fluorescence-dependent semi-quantitative method involving hybridization of unique barcodes to an array. We evaluated the method using three human breast cancer cell lines and identified specific gene expression profiles for each line. Furthermore, we applied the method to single cells and confirmed the heterogeneity of a cell population. Successful gene detection from cancer cells in human blood from metastatic breast cancer patients supports the use of RT-MLPA as a diagnostic tool for cancer genomics.

  • 15.
    Pradhananga, Sailendra
    et al.
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Spalinskas, Rapolas
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH).
    Poujade, Flore-Anne
    Karolinska Inst, Dept Med Solna, Ctr Mol Med, Cardiovasc Med Unit, Stockholm, Sweden.;Karolinska Univ Hosp, Solna, Sweden..
    Eriksson, Per
    Karolinska Inst, Dept Med Solna, Ctr Mol Med, Cardiovasc Med Unit, Stockholm, Sweden.;Karolinska Univ Hosp, Solna, Sweden..
    Sahlén, Pelin
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Promoter anchored interaction landscape of THP-1 macrophages captures early immune response processes2020In: Cellular Immunology, ISSN 0008-8749, E-ISSN 1090-2163, Vol. 355, article id 104148Article in journal (Refereed)
    Abstract [en]

    Macrophages are highly plastic immune cells with temporally distinct transcriptome changes upon lipopolysaccride (LPS) activation. However, to what extent transcriptome reprogramming is mediated via spatial chromatin looping is not well studied. We generated high resolution chromatin interaction maps for LPS-stimulated THP-1 macrophages (0 and 2 h) using capture Hi-C. Success of LPS stimulation was validated with transcriptome sequencing. Circa 2900 genes changed their interaction profile upon LPS stimulation and those gaining interactions were enriched for LPS response relevant processes, suggesting a substantial role for distal regulation. Immune and cardiovascular risk variants were enriched within the interacting regions, thereby providing insights into macrophage biology.

  • 16.
    Pradhananga, Sailendra
    et al.
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Spalinskas, Rapolas
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Poujade, Flore-Anne
    Karolinska Inst, Dept Med Solna, Ctr Mol Med, Cardiovasc Med Unit, Stockholm, Sweden.;Karolinska Univ Hosp, Solna, Sweden..
    Eriksson, Per
    Karolinska Inst, Dept Med Solna, Ctr Mol Med, Cardiovasc Med Unit, Stockholm, Sweden.;Karolinska Univ Hosp, Solna, Sweden..
    Sahlén, Pelin
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    The role of rare enhancer variants inbicuspid aortic valve pathologyManuscript (preprint) (Other academic)
    Abstract [en]

    Bicuspid aortic valve (BAV) is a heritable congenital valve defect associated with a multitude of heart complication. BAV is a highly heritable and relatively rare disease, however in most disease cases, no coding variant can be causally linked to the disease. Given the preponderance of heritability, we sought to understand the role of non-coding rare variants in bicuspid aortic valve pathology. To this end, we generated promoters-enhancer interaction maps (HiCap), transcriptome and H3K27Ac-enhancer profiles of aortic endothelial cells derived from individuals with bicuspid or tricuspid aortic valve. Further, we sequenced the entire genome of all individuals in our study and identified the rare variants (minor allele frequency < 0.5%). Using functional and context dependent datasets, we report three-fold enrichment of non-coding rare variants in enhancer regions. Moreover, the target genes of enhancers with rare variants were relevant for valve pathology only in BAV samples. This suggests that rare non-coding variants could have significant consequences for BAV pathology

  • 17.
    Sahlén, Pelin
    et al.
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Abdullayev, Ilgar
    Ramsköld, Daniel
    Matskova, Liudmila
    Rilakovic, Nemanja
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lötstedt, Britta
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Albert, Thomas J.
    Lundeberg, Joakim
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Sandberg, Rickard
    Genome-wide mapping of promoter-anchored interactions with close to single-enhancer resolution2015In: Genome Biology, ISSN 1465-6906, E-ISSN 1474-760X, Vol. 16, article id 156Article in journal (Refereed)
    Abstract [en]

    Although the locations of promoters and enhancers have been identified in several cell types, we still have limited information on their connectivity. We developed HiCap, which combines a 4-cutter restriction enzyme Hi-C with sequence capture of promoter regions. Applying the method to mouse embryonic stem cells, we identified promoter-anchored interactions involving 15,905 promoters and 71,984 distal regions. The distal regions were enriched for enhancer marks and transcription, and had a mean fragment size of only 699 bp - close to single-enhancer resolution. High-resolution maps of promoter-anchored interactions with HiCap will be important for detailed characterizations of chromatin interaction landscapes.

  • 18.
    Sahlén, Pelin
    et al.
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Spalinskas, Rapolas
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH).
    Asad, S.
    Mahapatra, K. D.
    Höjer, Pontus
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Anil, Anandashankar
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Eisfeldt, J.
    Srivastava, A.
    Nikamo, P.
    Mukherjee, A.
    Kim, K. -H
    Bergman, O.
    Ståhle, M.
    Sonkoly, E.
    Pivarcsi, A.
    Wahlgren, C. -F
    Nordenskjöld, M.
    Taylan, F.
    Bradley, M.
    Tapia-Páez, I.
    Chromatin interactions in differentiating keratinocytes reveal novel atopic dermatitis– and psoriasis-associated genes2020In: Journal of Allergy and Clinical Immunology, ISSN 0091-6749, E-ISSN 1097-6825Article in journal (Refereed)
    Abstract [en]

    Background: Hundreds of variants associated with atopic dermatitis (AD) and psoriasis, 2 common inflammatory skin disorders, have previously been discovered through genome-wide association studies (GWASs). The majority of these variants are in noncoding regions, and their target genes remain largely unclear. Objective: We sought to understand the effects of these noncoding variants on the development of AD and psoriasis by linking them to the genes that they regulate. Methods: We constructed genomic 3-dimensional maps of human keratinocytes during differentiation by using targeted chromosome conformation capture (Capture Hi-C) targeting more than 20,000 promoters and 214 GWAS variants and combined these data with transcriptome and epigenomic data sets. We validated our results with reporter assays, clustered regularly interspaced short palindromic repeats activation, and examination of patient gene expression from previous studies. Results: We identified 118 target genes of 82 AD and psoriasis GWAS variants. Differential expression of 58 of the 118 target genes (49%) occurred in either AD or psoriatic lesions, many of which were not previously linked to any skin disease. We highlighted the genes AFG1L, CLINT1, ADO, LINC00302, and RP1-140J1.1 and provided further evidence for their potential roles in AD and psoriasis. Conclusions: Our work focused on skin barrier pathology through investigation of the interaction profile of GWAS variants during keratinocyte differentiation. We have provided a catalogue of candidate genes that could modulate the risk of AD and psoriasis. Given that only 35% of the target genes are the gene nearest to the known GWAS variants, we expect that our work will contribute to the discovery of novel pathways involved in AD and psoriasis.

  • 19.
    Sahlén, Pelin
    et al.
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Yanhu, Liu
    Xu, Jinrui
    Yale Univ, Program Computat Biol & Bioinformat, New Haven, CT USA..
    Kubinyi, Eniko
    Eotvos Lorand Univ, Dept Ethol, Budapest, Hungary..
    Wang, Guo-Dong
    Chinese Acad Sci, Kunming Inst Zool, State Key Lab Genet Resources & Evolut, Kunming, Yunnan, Peoples R China.;Chinese Acad Sci, Ctr Excellence Anim Evolut & Genet, Kunming, Yunnan, Peoples R China..
    Savolainen, Peter
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Variants That Differentiate Wolf and Dog Populations Are Enriched in Regulatory Elements2021In: Genome Biology and Evolution, ISSN 1759-6653, E-ISSN 1759-6653, Vol. 13, no 4, article id evab076Article in journal (Refereed)
    Abstract [en]

    Research on the genetics of domestication most often focuses on the protein-coding exons. However, exons cover only a minor part (1-2%) of the canine genome, whereas functional mutations may be located also in regions beyond the exome, in regulatory regions. Therefore, a large proportion of phenotypical differences between dogs and wolves may remain genetically unexplained. In this study, we identified variants that have high allelic frequency differences (i.e., highly differentiated variants) between wolves and dogs across the canine genome and investigated the potential functionality. We found that the enrichment of highly differentiated variants was substantially higher in promoters than in exons and that such variants were enriched also in enhancers. Several enriched pathways were identified including oxytocin signaling, carbohydrate digestion and absorption, cancer risk, and facial and body features, many of which reflect phenotypes of potential importance during domestication, including phenotypes of the domestication syndrome. The results highlight the importance of regulatory mutations during dog domestication and motivate the functional annotation of the noncoding part of the canine genome.

  • 20. Shirley, B. C.
    et al.
    Mucaki, E. J.
    Whitehead, T.
    Costea, Paul Igor
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Akan, Pelin
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Rogan, P. K.
    Interpretation, Stratification and Evidence for Sequence Variants Affecting mRNA Splicing in Complete Human Genome Sequences2013In: Genomics, Proteomics and Bioinformatics, ISSN 1672-0229, Vol. 11, no 2, p. 77-85Article in journal (Refereed)
    Abstract [en]

    Information theory-based methods have been shown to be sensitive and specific for predicting and quantifying the effects of non-coding mutations in Mendelian diseases. We present the Shannon pipeline software for genome-scale mutation analysis and provide evidence that the software predicts variants affecting mRNA splicing. Individual information contents (in bits) of reference and variant splice sites are compared and significant differences are annotated and prioritized. The software has been implemented for CLC-Bio Genomics platform. Annotation indicates the context of novel mutations as well as common and rare SNPs with splicing effects. Potential natural and cryptic mRNA splicing variants are identified, and null mutations are distinguished from leaky mutations. Mutations and rare SNPs were predicted in genomes of three cancer cell lines (U2OS, U251 and A431), which were supported by expression analyses. After filtering, tractable numbers of potentially deleterious variants are predicted by the software, suitable for further laboratory investigation. In these cell lines, novel functional variants comprised 6-17 inactivating mutations, 1-5 leaky mutations and 6-13 cryptic splicing mutations. Predicted effects were validated by RNA-seq analysis of the three aforementioned cancer cell lines, and expression microarray analysis of SNPs in HapMap cell lines.

  • 21.
    Solnestam, Beata W.
    et al.
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Kvastad, Linda
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Johansson, Elin
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Nygren, A. O.
    Laddach, N.
    Sahlén, Pellin
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Vickovic, Sanja
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lundberg, Joakim
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Single cell analysis of cancer cells using an improved RT-MLPA method has potential for cancer diagnosis and monitoringManuscript (preprint) (Other academic)
  • 22.
    Stranneheim, Henrik
    et al.
    KTH, School of Biotechnology (BIO), Gene Technology.
    Werne Solnestam, Beata
    KTH, School of Biotechnology (BIO), Gene Technology.
    Akan, Pelin
    KTH, School of Biotechnology (BIO), Gene Technology.
    Lundberg, Emma
    KTH, School of Biotechnology (BIO), Proteomics.
    Lundeberg, Joakim
    KTH, School of Biotechnology (BIO), Gene Technology.
    Transcript nuclear retention effects quantification of gene expression levelsManuscript (preprint) (Other academic)
    Abstract [en]

    The majority of published differential gene expression studies have used RNA isolated from whole cell extracts (total RNA), overlooking the potential impact of including the nuclear transcriptome in the analyses. It has not been firmly established that the contribution of nuclear RNA is negligible or how the inclusion of it affects quantification of gene expression. Previous studies have estimated that the nuclear transcriptome is five to ten times more complex than the cytoplasmic [1]. Hence, RNA purified solely from the cytoplasm should have fewer unique transcripts, resulting in more sequence counts per transcript and resulting in increased power to detect remaining transcripts. In this study, cytoplasmic and total mRNA have been prepared from three human cell‐lines and sequenced using massive parallel sequencing. The resulting sequence data was analyzed regarding the effect of number of biological replicates, read length and transcripts fractionation on calling differentially detected genes. In addition, the impact of length and secondary structure of mRNAs un‐translated regions (UTRs), and coding sequence length on nucleus to cytoplasm transportation rates of mRNAs was studied. We observe that the number of differentially detected genes was not significantly increased by adding more than three biological replicates or by increasing the sequence read length > 35bp. More differentially detected genes were found in the cytoplasmic RNA compared to the total RNA and a nuclear retention effect was observed for transcripts with long and structured 5’‐ and 3’‐UTR or long protein coding sequences.

  • 23.
    Ståhl, Patrik
    et al.
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Salmén, Fredrik
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Vickovic, Sanja
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lundmark, Anna
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Fernandez Navarro, Jose
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Magnusson, Jens
    Giacomello, Stefania
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Asp, Michaela
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Westholm, Jakub O.
    Huss, Mikael
    Mollbrink, Annelie
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Linnarsson, Sten
    Codeluppi, Simone
    Borg, Ake
    Ponten, Fredrik
    Costea, Paul Igor
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Biotechnology (BIO), Gene Technology.
    Sahlén, Pelin
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Mulder, Jan
    Bergmann, Olaf
    Lundeberg, Joakim
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Frisen, Jonas
    Visualization and analysis of gene expression in tissue sections by spatial transcriptomics2016In: Science, ISSN 0036-8075, E-ISSN 1095-9203, Vol. 353, no 6294, p. 78-82Article in journal (Refereed)
    Abstract [en]

    Analysis of the pattern of proteins or messenger RNAs (mRNAs) in histological tissue sections is a cornerstone in biomedical research and diagnostics. This typically involves the visualization of a few proteins or expressed genes at a time. We have devised a strategy, which we call "spatial transcriptomics," that allows visualization and quantitative analysis of the transcriptome with spatial resolution in individual tissue sections. By positioning histological sections on arrayed reverse transcription primers with unique positional barcodes, we demonstrate high-quality RNA-sequencing data with maintained two-dimensional positional information from the mouse brain and human breast cancer. Spatial transcriptomics provides quantitative gene expression data and visualization of the distribution of mRNAs within tissue sections and enables novel types of bioinformatics analyses, valuable in research and diagnostics.

  • 24. Sánchez, J. L. A.
    et al.
    Henry, O. Y. F.
    Joda, H.
    Solnestam, Beata Werne
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Kvastad, Linda
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Johansson, Elin
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Akan, Pelin
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lundeberg, Joakim
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lladach, N.
    Ramakrishnan, D.
    Riley, I.
    O'Sullivan, C. K.
    Multiplex PCB-based electrochemical detection of cancer biomarkers using MLPA-barcode approach2016In: Biosensors & bioelectronics, ISSN 0956-5663, E-ISSN 1873-4235, Vol. 82, p. 224-232Article in journal (Refereed)
    Abstract [en]

    Asymmetric multiplex ligation-dependent probe amplification (MLPA) was developed for the amplification of seven breast cancer related mRNA markers and the MLPA products were electrochemically detected via hybridization. Seven breast cancer genetic markers were amplified by means of the MLPA reaction, which allows for multiplex amplification of multiple targets with a single primer pair. Novel synthetic MLPA probes were designed to include a unique barcode sequence in each amplified gene. Capture probes complementary to each of the barcode sequences were immobilized on each electrode of a low-cost electrode microarray manufactured on standard printed circuit board (PCB) substrates. The functionalised electrodes were exposed to the single-stranded MLPA products and following hybridization, a horseradish peroxidase (HRP)-labelled DNA secondary probe complementary to the amplified strand completed the genocomplex, which was electrochemically detected following substrate addition. The electrode arrays fabricated using PCB technology exhibited an excellent electrochemical performance, equivalent to planar photolithographically-fabricated gold electrodes, but at a vastly reduced cost (>50 times lower per array). The optimised system was demonstrated to be highly specific with negligible cross-reactivity allowing the simultaneous detection of the seven mRNA markers, with limits of detections as low as 25 pM. This approach provides a novel strategy for the genetic profiling of tumour cells via integrated "amplification-to-detection".

  • 25. Tapia-Paez, I.
    et al.
    Asad, S.
    Taylan, F.
    Spalinskas, Rapolas
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Anandashankar, A.
    Nordenskjold, M.
    Wahlgren, C. F.
    Sahlén, Pelin
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Bradley, M.
    Studies of keratinocyte-specific regulatory interactions by three-dimensional mapping with a focus on atopic dermatitis2018In: British Journal of Dermatology, ISSN 0007-0963, E-ISSN 1365-2133, Vol. 179, no 1, p. E33-E33Article in journal (Refereed)
  • 26.
    Wang, Sailan
    et al.
    Karolinska Inst, Dept Med, Solna, Stockholm, Sweden..
    Vaz, Raquel
    Karolinska Inst, Dept Mol Med & Surg, Stockholm, Sweden..
    Sahlén, Pelin
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Wahlgren, Carl-Fredrik
    Karolinska Inst, Dept Med, Solna, Stockholm, Sweden..
    Nordenskjold, Magnus
    Karolinska Inst, Dept Med, Solna, Stockholm, Sweden..
    Tapia-Paez, Isabel
    Karolinska Inst, Dept Med, Solna, Stockholm, Sweden..
    Bradley, Maria
    Karolinska Inst, Dept Med, Solna, Stockholm, Sweden..
    Exploring the Role of the Ado Gene in Atopic Dermatitis2023In: Acta Dermato-Venereologica, ISSN 0001-5555, E-ISSN 1651-2057, Vol. 103, p. 42-43Article in journal (Other academic)
  • 27.
    Werne Solnestam, Beata
    et al.
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Stranneheim, Henrik
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Hällman, Jimmie
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Käller, Max
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lundberg, Emma
    KTH, School of Biotechnology (BIO), Proteomics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lundeberg, Joakim
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Akan, Pelin
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Comparison of total and cytoplasmic mRNA reveals global regulation by nuclear retention and miRNAs2012In: BMC Genomics, E-ISSN 1471-2164, Vol. 13, no 1, p. 574-Article in journal (Refereed)
    Abstract [en]

    Background: The majority of published gene-expression studies have used RNA isolated from whole cells, overlooking the potential impact of including nuclear transcriptome in the analyses. In this study, mRNA fractions from the cytoplasm and from whole cells (total RNA) were prepared from three human cell lines and sequenced using massive parallel sequencing. Results: For all three cell lines, of about 15000 detected genes approximately 400 to 1400 genes were detected in different amounts in the cytoplasmic and total RNA fractions. Transcripts detected at higher levels in the total RNA fraction had longer coding sequences and higher number of miRNA target sites. Transcripts detected at higher levels in the cytoplasmic fraction were shorter or contained shorter untranslated regions. Nuclear retention of transcripts and mRNA degradation via miRNA pathway might contribute to this differential detection of genes. The consequence of the differential detection was further investigated by comparison to proteomics data. Interestingly, the expression profiles of cytoplasmic and total RNA correlated equally well with protein abundance levels indicating regulation at a higher level. Conclusions: We conclude that expression levels derived from the total RNA fraction be regarded as an appropriate estimate of the amount of mRNAs present in a given cell population, independent of the coding sequence length or UTRs.

  • 28.
    Zaugg, Judith Barbara
    et al.
    European Molecular Biology Laboratory, Heidelberg, Germany.
    Sahlén, Pelin
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Andersson, Robin
    Section for Computational and RNA Biology, Department of Biology, University of Copenhagen, Copenhagen, Denmark; The Novo Nordisk Foundation Center for Genomic Mechanisms of Disease, Broad Institute of MIT and Harvard, Cambridge, MA, USA.
    Alberich-Jorda, Meritxell
    Department of Hemato-oncology, Institute of Molecular Genetics of the CAS, Prague, Czech Republic; Childhood Leukaemia Investigation Prague, Department of Pediatric Haematology and Oncology, 2nd Faculty of Medicine, Charles University in Prague, University Hospital Motol, Prague, Czech Republic.
    de Laat, Wouter
    Hubrecht Institute-KNAW, Oncode Institute and University Medical Center Utrecht, Utrech, Netherlands.
    Deplancke, Bart
    Laboratory of Systems Biology and Genetics, Institute of Bio-engineering, School of Life Sciences, EPFL, Lausanne, Switzerland.
    Ferrer, Jorge
    Centre for Genomic Regulation, the Barcelona Institute of Science and Technology, Barcelona, Spain; Centro de Investigación Biomédica en red Diabetes y enfermedades metabólicas asociadas (CIBERDEM), Madrid, Spain; Department of Metabolism, Digestion and Reproduction, Imperial College London, London, UK.
    Mandrup, Susanne
    The Novo Nordisk Foundation Center for Genomic Mechanisms of Disease, Broad Institute of MIT and Harvard, Cambridge, MA, USA; Functional Genomics and Metabolism Research Unit, Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense, Denmark.
    Natoli, Gioacchino
    IEO, European Institute of Oncology – IRCCS, Milan, Italy.
    Plewczynski, Dariusz
    Laboratory of Functional and Structural Genomics, Centre of New Technologies, University of Warsaw, Warsaw, Poland.
    Rada-Iglesias, Alvaro
    Institute of Biomedicine and Biotechnology of Cantabria (IBBTEC), CSIC/University of Cantabria, Santander, Spain.
    Spicuglia, Salvatore
    TAGC, UMR 1090, Aix-Marseille University, INSERM, Marseille, France; Equipe Labellisée, LIGUE Contre le Cancer, Marseille, France.
    Current challenges in understanding the role of enhancers in disease2022In: Nature Structural & Molecular Biology, ISSN 1545-9993, E-ISSN 1545-9985, Vol. 29, no 12, p. 1148-1158Article in journal (Refereed)
    Abstract [en]

    Enhancers play a central role in the spatiotemporal control of gene expression and tend to work in a cell-type-specific manner. In addition, they are suggested to be major contributors to phenotypic variation, evolution and disease. There is growing evidence that enhancer dysfunction due to genetic, structural or epigenetic mechanisms contributes to a broad range of human diseases referred to as enhanceropathies. Such mechanisms often underlie the susceptibility to common diseases, but can also play a direct causal role in cancer or Mendelian diseases. Despite the recent gain of insights into enhancer biology and function, we still have a limited ability to predict how enhancer dysfunction impacts gene expression. Here we discuss the major challenges that need to be overcome when studying the role of enhancers in disease etiology and highlight opportunities and directions for future studies, aiming to disentangle the molecular basis of enhanceropathies.

  • 29.
    Zhang, Miao
    et al.
    KTH, School of Information and Communication Technology (ICT), Materials- and Nano Physics, Material Physics, MF.
    Schmidt, Torsten
    KTH, School of Information and Communication Technology (ICT), Materials- and Nano Physics, Material Physics, MF.
    Jemt, Anders
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Sahlén, Pelin
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Sychugov, Ilya
    KTH, School of Information and Communication Technology (ICT), Materials- and Nano Physics, Material Physics, MF.
    Lundeberg, Joakim
    KTH, School of Biotechnology (BIO), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Linnros, Jan
    KTH, School of Information and Communication Technology (ICT), Materials- and Nano Physics, Material Physics, MF.
    Nanopore arrays in a silicon membrane for parallel single-molecule detection: DNA translocation2015In: Nanotechnology, ISSN 0957-4484, E-ISSN 1361-6528, Vol. 26, no 31, article id 314002Article in journal (Refereed)
    Abstract [en]

    Optical nanopore sensing offers great potential in single-molecule detection, genotyping, or DNA sequencing for high-throughput applications. However, one of the bottle-necks for fluorophore-based biomolecule sensing is the lack of an optically optimized membrane with a large array of nanopores, which has large pore-to-pore distance, small variation in pore size and low background photoluminescence (PL). Here, we demonstrate parallel detection of single-fluorophore-labeled DNA strands (450 bps) translocating through an array of silicon nanopores that fulfills the above-mentioned requirements for optical sensing. The nanopore array was fabricated using electron beam lithography and anisotropic etching followed by electrochemical etching resulting in pore diameters down to similar to 7 nm. The DNA translocation measurements were performed in a conventional wide-field microscope tailored for effective background PL control. The individual nanopore diameter was found to have a substantial effect on the translocation velocity, where smaller openings slow the translocation enough for the event to be clearly detectable in the fluorescence. Our results demonstrate that a uniform silicon nanopore array combined with wide-field optical detection is a promising alternative with which to realize massively-parallel single-molecule detection.

  • 30.
    Zhang, Wensheng
    et al.
    Soochow Univ, Cam Su Genom Resource Ctr, Suzhou 215123, Peoples R China.;Wellcome Sanger Inst, Hinxton CB10 1SA, England..
    Chronis, Constantinos
    Univ Calif Los Angeles, David Geffen Sch Med, Dept Biol & Chem, Los Angeles, CA 90095 USA.;Univ Calif Los Angeles, Eli & Edythe Broad Ctr Regenerat Med & Stem Cell, Los Angeles, CA USA.;Univ Calif Los Angeles, Jonsson Comprehens Canc Ctr, Bioinformat Program, Los Angeles, CA 90024 USA.;Univ Calif Los Angeles, Mol Biol Inst, Los Angeles, CA 90095 USA..
    Chen, Xi
    Wellcome Sanger Inst, Hinxton CB10 1SA, England..
    Zhang, Heyao
    Soochow Univ, Cam Su Genom Resource Ctr, Suzhou 215123, Peoples R China..
    Spalinskas, Rapolas
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Pardo, Mercedes
    Chester Beatty Labs, Inst Canc Res, London, England..
    Chen, Liangliang
    Soochow Univ, Cam Su Genom Resource Ctr, Suzhou 215123, Peoples R China..
    Wu, Guangming
    Max Planck Inst Mol Biomed, Dept Cell & Dev Biol, Rontgenstr 20, D-48149 Munster, Germany..
    Zhu, Zhexin
    Wellcome Sanger Inst, Hinxton CB10 1SA, England..
    Yu, Yong
    Wellcome Sanger Inst, Hinxton CB10 1SA, England..
    Yu, Lu
    Chester Beatty Labs, Inst Canc Res, London, England..
    Choudhary, Jyoti
    Chester Beatty Labs, Inst Canc Res, London, England..
    Nichols, Jennifer
    Univ Cambridge, Wellcome Trust Med Res Council, Stem Cell Inst, Tennis Court Rd, Cambridge CB2 1QR, England..
    Parast, Mana M.
    Univ Calif San Diego, Dept Pathol, La Jolla, CA 92093 USA.;Univ Calif San Diego, Sanford Consortium Regenerat Med, La Jolla, CA 92093 USA..
    Greber, Boris
    Max Planck Inst Mol Biomed, Dept Cell & Dev Biol, Rontgenstr 20, D-48149 Munster, Germany..
    Sahlén, Pelin
    KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Plath, Kathrin
    Univ Calif Los Angeles, David Geffen Sch Med, Dept Biol & Chem, Los Angeles, CA 90095 USA.;Univ Calif Los Angeles, Eli & Edythe Broad Ctr Regenerat Med & Stem Cell, Los Angeles, CA USA.;Univ Calif Los Angeles, Jonsson Comprehens Canc Ctr, Bioinformat Program, Los Angeles, CA 90024 USA.;Univ Calif Los Angeles, Mol Biol Inst, Los Angeles, CA 90095 USA..
    The BAF and PRC2 Complex Subunits Dpf2 and Eed Antagonistically Converge on Tbx3 to Control ESC Differentiation2019In: Cell Stem Cell, ISSN 1934-5909, E-ISSN 1875-9777, Vol. 24, no 1, p. 138-+Article in journal (Refereed)
    Abstract [en]

    BAF complexes are composed of different subunits with varying functional and developmental roles, although many subunits have not been examined in depth. Here we show that the Baf45 subunit Dpf2 maintains pluripotency and ESC differentiation potential. Dpf2 co-occupies enhancers with Oct4, Sox2, p300, and the BAF subunit Brg1, and deleting Dpf2 perturbs ESC self-renewal, induces repression of Tbx3, and impairs mesendodermal differentiation without dramatically altering Brg1 localization. Mesendodermal differentiation can be rescued by restoring Tbx3 expression, whose distal enhancer is positively regulated by Dpf2-dependent H3K27ac maintenance and recruitment of pluripotency TFs and Brg1. In contrast, the PRC2 subunit Eed binds an intragenic Tbx3 enhancer to oppose Dpf2-dependent Tbx3 expression and mesendodermal differentiation. The PRC2 subunit Ezh2 likewise opposes Dpf2-dependent differentiation through a distinct mechanism involving Nanog repression. Together, these findings delineate distinct mechanistic roles for specific BAF and PRC2 subunits during ESC differentiation.

  • 31.
    Zhigulev, Artemii
    et al.
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Lang, K.
    Karolinska Inst, Cardiovasc Med Unit, Dept Med Solna, Karolinska Univ Hosp, Stockholm, Sweden..
    Pradhananga, Sailendra
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Spalinskas, Rapolas
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Franco-Cereceda, A.
    Karolinska Inst, Dept Mol Med & Surg, Karolinska Univ Hosp, Stockholm, Sweden..
    Bjorck, H.
    Karolinska Inst, Cardiovasc Med Unit, Dept Med Solna, Karolinska Univ Hosp, Stockholm, Sweden..
    Eriksson, P.
    Karolinska Inst, Cardiovasc Med Unit, Dept Med Solna, Karolinska Univ Hosp, Stockholm, Sweden..
    Sahlén, Pelin
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    The Role Of Rare Enhancer Variants In Bicuspid Aortic Valve Pathology2022In: Atherosclerosis, ISSN 0021-9150, E-ISSN 1879-1484, Vol. 355, p. E68-E69Article in journal (Other academic)
  • 32.
    Zhigulev, Artemii
    et al.
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Norberg, Zandra
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Cordier, Julie
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Spalinskas, Rapolas
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Bassereh, Hassan
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Björn, Niclas
    Division of Clinical Chemistry and Pharmacology, Department of Biomedical and Clinical Sciences, Faculty of Medicine and Health Sciences, Linköping University, Linköping, Sweden.
    Pradhananga, Sailendra
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Gréen, Henrik
    Division of Clinical Chemistry and Pharmacology, Department of Biomedical and Clinical Sciences, Faculty of Medicine and Health Sciences, Linköping University, Linköping, Sweden;Department of Forensic Genetics and Forensic Toxicology, National Board of Forensic Medicine, Linköping, Sweden.
    Sahlén, Pelin
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Enhancer mutations modulate the severity of chemotherapy-induced myelosuppression2024In: Life Science Alliance, E-ISSN 2575-1077, Vol. 7, no 3, p. e202302244-e202302244Article in journal (Refereed)
    Abstract [en]

    Non-small cell lung cancer is often diagnosed at advanced stages, and many patients are still treated with classical chemotherapy. The unselective nature of chemotherapy often results in severe myelosuppression. Previous studies showed that protein-coding mutations could not fully explain the predisposition to myelosuppression. Here, we investigate the possible role of enhancer mutations in myelosuppression susceptibility. We produced transcriptome and promoter-interaction maps (using HiCap) of three blood stem-like cell lines treated with carboplatin or gemcitabine. Taking advantage of publicly available enhancer datasets, we validated HiCap results in silico and in living cells using epigenetic CRISPR technology. We also developed a network approach for interactome analysis and detection of differentially interacting genes. Differential interaction analysis provided additional information on relevant genes and pathways for myelosuppression compared with differential gene expression analysis at the bulk level. Moreover, we showed that enhancers of differentially interacting genes are highly enriched for variants associated with differing levels of myelosuppression. Altogether, our work represents a prominent example of integrative transcriptome and gene regulatory datasets analysis for the functional annotation of noncoding mutations.

  • 33.
    Zhigulev, Artemii
    et al.
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Sahlén, Pelin
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology.
    Targeted Chromosome Conformation Capture (HiCap)2022In: Spatial Genome Organization, Springer Nature , 2022, p. 75-94Chapter in book (Refereed)
    Abstract [en]

    Targeted chromosome conformation capture (HiCap) is an experimental method for detecting spatial interactions of genomic features such as promoters and/or enhancers. The protocol first describes the design of sequence capture probes. After that, it provides details on the chromosome conformation capture adapted for next-generation sequencing (Hi-C). Finally, the methodology for coupling Hi-C with sequence capture technology is described. 

  • 34.
    Åkerborg, Örjan
    et al.
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Spalinskas, Rapolas
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Pradhananga, Sailendra
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Anil, Anandashankar
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Höjer, Pontus
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Poujade, Flore-Anne
    Karolinska Inst, Cardiovasc Med Unit, Dept Med, Ctr Mol Med, Stockholm, Sweden..
    Folkersen, Lasse
    Tech Univ Denmark, Dept Bioinformat, Copenhagen, Denmark..
    Sahlén, Pelin
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Gene Technology. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Eriksson, Per
    Karolinska Inst, Cardiovasc Med Unit, Dept Med, Ctr Mol Med, Stockholm, Sweden..
    High-Resolution Regulatory Maps Connect Vascular Risk Variants to Disease-Related Pathways2019In: Circulation: Genomic and Precision Medicine, E-ISSN 2574-8300 , Vol. 12, no 3, article id e002353Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Genetic variant landscape of coronary artery disease is dominated by noncoding variants among which many occur within putative enhancers regulating the expression levels of relevant genes. It is crucial to assign the genetic variants to their correct genes both to gain insights into perturbed functions and better assess the risk of disease. METHODS: In this study, we generated high-resolution genomic interaction maps (similar to 750 bases) in aortic endothelial, smooth muscle cells and THP-1 (human leukemia monocytic cell line) macrophages stimulated with lipopolysaccharide using Hi-C coupled with sequence capture targeting 25 429 features, including variants associated with coronary artery disease. We also sequenced their transcriptomes and mapped putative enhancers using chromatin immunoprecipitation with an antibody against H3K27Ac. RESULTS: The regions interacting with promoters showed strong enrichment for enhancer elements and validated several previously known interactions and enhancers. We detected interactions for 727 risk variants obtained by genome-wide association studies and identified novel, as well as established genes and functions associated with cardiovascular diseases. We were able to assign potential target genes for additional 398 genome-wide association studies variants using haplotype information, thereby identifying additional relevant genes and functions. Importantly, we discovered that a subset of risk variants interact with multiple promoters and their expression levels were strongly correlated. CONCLUSIONS: In summary, we present a catalog of candidate genes regulated by coronary artery disease-related variants and think that it will be an invaluable resource to further the investigation of cardiovascular pathologies and disease.

1 - 34 of 34
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