Has anyone used this attached protocol before? It seems easy but they have only used that for sheep blood. Do you think it would work with human blood as they have suggested?
Dear Harindra
Please accept this book (PCR Protocols) as a gift from me and read chapter 6 for a detailed look at manual procedure for extracting DNA from Whole Blood.
I am sure that this book will solve many other problems for you also that may arise during your future research.
Best Regards
Look for Erythrocytes lysis buffer. This described method is difficult, old and inefficient
Dear Elena, do you have any suggestions? or a simple protocol that I can use?
With Human Blood I have very good experience with Qiagen Kit, but also from Macherey-Nagel or Peqlab a good kit is available. From the latter company you normally can easily get some sample kit for 5-10 extractions, just ask them...
G'day Harindra, the protocol will probably work but the DNA might not be as pure as DNA extracted using a commercial DNA extraction kit. I've had success with the Purelink Genomic DNA Mini Kit from LifeTechnologies. Do you want RNA as well?
Thanks, I would only need DNA from these blood samples. I may also use a commercial kit (Qiagen preferably) in parallel. But this is just to do the same extraction manually. So I can probably compare which one is the best.
Normally the quality in between different kits of different companies doesn't differ that much, but kits are normally faster and quality of DNA / RNA is most times better.
Hi
Take look at this article:
https://www.researchgate.net/publication/13582263_Optimized_procedure_for_DNA_isolation_from_fresh_and_cryopreserved_clotted_human_blood_useful_in_clinical_molecular_testing?ev=prf_pub
Article Optimized procedure for DNA isolation from fresh and cryopre...
@ Ted: I was using Qiagen Midi kit for 2 ml of blood and I isolated DNA of several hundret samples. The quality was good, A260/280 value was always coming in the range of 1.8+-0.5 and the amound of DNA was also very consistent in a good yield. This blood was frozen before as it came from a tumor bank for up to 2 years...
I am working on Rheumatoid arthritis (RA) patients and SNP analysis, we are using extracting DNA from Human blood almost daily. Instead of kit we are using manual method for DNA isolation. So a basic protocol which we are following attached here and I am glad to share we have isolated highly pure DNA each time.
It may help you
Dear Harindra,
If funds are restriction - you may continue using manual method which works very well still and in such case there is no need to compare it against quality of DNA using Kits.
But, same time I support Mr. Ted Michelini and Mr. Mahendra Verma that the shared protocol will not disappoint you either and with some initial optimization in your hands will still give quite good quality yield. Only difference would be number of extractions Vs time.
Before the Kits came into existence & luxury of funds, way back in early 90's we were using manual protocols with acceptable quality. Protocol shared by MKV should definitely work, partly dependent on experimenter's hands.
Best wishes,
Thank you Sanjeev, appreciate your explanation. It is like I'm doing this study for 2 different bodies. And I'll probably get adequate funds for a KIT from one body so I'll always have a backup if the manual extraction doesn't work for the other body. Just wanted to make sure that shared protocol will work because it seems easy and doesn't require whole lots of reagents. My only concern was authors had used that for sheeps but not for human. But they had suggested that this protocol should work in humans as well. Thanks
Dear Harindra
Please accept this book (PCR Protocols) as a gift from me and read chapter 6 for a detailed look at manual procedure for extracting DNA from Whole Blood.
I am sure that this book will solve many other problems for you also that may arise during your future research.
Best Regards
A company called KAPA has a Polymerase that they claim can be used to amplify DNA from whole blood. It is not my field of expertise, so I have never used their kit and do not know if it works or not, but might be worth a shot for people who do that routinely.
http://www.kapabiosystems.com/products/name/kapa-blood-pcr-kits
hello. can any help me.
i am a tyro researcher. want to work on oral cancer OSF at genetic level for early detection of ca. here are no.of genes worked by researcher. i need to work in my population but donot know which SNP's should i concentrate on and how can i find novelity. thanks
hi every body.
Someone can introduce a valid protocol referances without the need for a kit to extract DNA or RNA? thanks.
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