Evaluation of methods for isolating cell-free DNA in blood
Independent thesis Advanced level (degree of Master (Two Years)), 20 credits / 30 HE creditsStudent thesis
The presence of cell-free DNA (cfDNA) in blood has been known for some time but it was just recently that certain applications for it materialized. First it was shown that elevated levels of cell-free DNA in cancer patients had the potential of serving as a biomarker and later, fetal DNA was found in cfDNA in maternal blood and could therefore be used for non-invasive prenatal diagnostics. Several other diseases such a s heart attack and tissue injures after stroke and trauma have also showed increased levels of cfDNA. The concentration of cfDNA in blood is often low and varies between individuals, which requires reliable and efficient methods for isolation of the DNA before it can be used for diagnosis. Three isolation methods were evaluated in this work and compared in terms of yield and the potential of becoming automated. All methods were based on the same principle where cfDNA binds to a solid support, either a filter or magnetic beads, when right bindingconditions are obtained. We found that all three methods were capable of isolating cfDNA with comparable yields. In addition, although all methods use Proteinase K in the original lysis step, we found thta one of the methods managed without Proteinase K, which in combination with the discovery that the same method also worked with plasma centrifuged only once (instead of tow times that is standard). These findings are promising when considering an automated cfDNA isolation procedure, as plasma transfer steps and enzymatic incubations are eliminated.
Place, publisher, year, edition, pages
DNA, cell-free blood isolation
Engineering and Technology
IdentifiersURN: urn:nbn:se:kth:diva-173628OAI: oai:DiVA.org:kth-173628DiVA: diva2:854043