In the course of the most recent couple of decades, PCR, next generation sequencing and microarray advancements have taken blood-based research to another dimension. Modern blood-based application ranges from DNA fingerprinting, whole genome sequencing, blood banking to fluid biopsy and many more. Regardless of the application, pure, intact, double stranded and highly concentrated DNA extraction from whole blood is an essential prerequisite to success in this area. The technique chosen for whole blood DNA isolation impacts not only on the results but also on the ease of molecular biology workflow. As with many other scientific techniques, choosing an extraction method can be a challenge in itself. Laboratories generally have their own preferred tried and tested protocol. Protocols can differ regarding setup and the exact chemical substances used to lyse cells and selectively isolate DNA. However, almost all of the available protocols for whole blood DNA isolation involve these common steps: cell lysis, removal of protein, other contaminants and RNA, and (finally) DNA recovery. The genomic DNA Isolation from Blood is being carried out in the ASRBC LAB, ACI LTD. for various tests and as well as research purposes. The protocol followed in your lab is given in a flow chart which is as follows: 1) Blood Samples were collected and stored in EDTA vacutainer tubes frozen at -70°C 2) Buffer was mixed and then centrifuged 3) Supernatant removed and discarded 4) Step 2 & 3 are repeated 5) Sodium Acetate added to the pellets. SDS & Proteinase K added and after vortex it was incubated for 1 hour at 55°C 6) Phenol Isoamyl Alcohol was added and centrifuged 7) Aqueous layer removed. Add ethanol and incubate for 15 minutes at -20°C 8) Again centrifuge and discard the supernatant 9) TE Buffer was added and incubated for 10 minutes at 55°C 10) Add sodium acetate and ethanol and then centrifuge 11) Supernatant discarded and rinsed with ethanol. Then centrifugation was done 12) After discarding supernatant the pellets were dried 13) Dissolve the pellets in TE Buffer
The assistance of Research Assistant Aqief Afzal of ASRBC, ACI Ltd. is highly appreciated.