Tail DNA extraction for regular genotyping

Reagents

1. Cut tails from animals of 0.5 cm.

2. Prepare tail lysis buffer by adding proteinase K to the Viagen Lysis Reagent. For each tail, I use 250 ul lysis buffer which contains 2.5 ul Proteinase K solution in 247.5 ul Viagen Lysis Reagent.

3. Incubate at 55 C overnight. I usually put all the tubes into the hybridization glass bottle and keep rotating in the Isotemp incubator.

4. The next morning, vortex the tubes to mix the solution and boil it at 85 C for 45 min to inactivate Proteinase K.

5. Spin down at 14k RPM for 3 min.

6. Take 2 ul for genotyping.

7. This DNA preparation is very stable at RT. I have tested it after 1 week which does not affect the quality for genotyping.


Comments

Add a comment
Posted by ebiomethods Mon, 22 Dec 2008 22:02:38
Subject: Comments are welcome
Please give us your feedback to help us improve.
Posted by neverthink (nevernetbug) Tue, 23 Dec 2008 15:40:56
Subject: Another simpler way to get DNA
it would only take Alkaline lysis buffer 15-30 min at 95 degree and then neuralize with Acedic buffer and then go on with PCR, without doing any other steps. one could finish genotyping within 2-3hrs from cutting tail with minimum handling. This is now a very widely used protocol, also commercial kits available but not worth buying though.

Posted by guest Tue, 10 Feb 2009 08:09:57
Subject: please tell us the detailed components of Alkaline lysis and

Posted by ebiomethods Tue, 10 Mar 2009 16:09:34
Subject: reply to guest
Please refer to \\\'Tail DNA extraction for regular genotyping - an alternative inexpensive method\\\' for details.
Site News
New feature:
You can add comments on protocols now. Just go to any individual method page and click on the "Add a comment" link.

Highlight of the month
SIRT6 links glucose and lipid metabolism

SIRT1, a histone deacetylase, has drawn considerable attention due to its important roles in metabolic regulation and longevity. SIRT1 is induced by fasting and suppressed by feeding. SIRT1 increases glucose production and beta oxidation in liver to meet energy needs during food deprivation. Another member of SIRT family, SIRT6 follows similar expression patterns as SIRT1. The function of SIRT6 in metabolic regulation remains unknown.

Kim and colleagues found SIRT1 can regulate SIRT6 expression in liver. Specifically, SIRT1 functions together with FOXO3a and stimulates SIRT6 transcription during fasting. The activation of SIRT6 directly suppresses gene expression of metabolic enzymes involved in triglyceride synthesis and glycolysis in liver. Deficiency of SIRT6 causes increases of glucose utilization, reduced beta oxidation and as a result, fatty liver. More importantly, Kim et al found the expression of SIRT6 is decreased in human fatty liver samples which indicates SIRT6 may play critical roles in liver steatosis in clinical settings. This study landed in Cell Metabolism of this month. ... Read more highlights.