How to Isolate DNA from marine nematodes? Because marine nematodes are a microscopic organisms. How is it possible to isolate the single species and how to isolate DNA?
I advice you go through the website of QIAGEN company
http://www.facebook.com/QIAGENscience
If you have a handful of soil, sediments, organic matters, infested plant parts etc., you may extract those tiny organism following Cobb's decanting and sieving followed by modified Baermann's technique.
What do you want to do with the isolated DNA?
You could try the same method we use to isolate DNA from tissue samples for genotyping:
DNA isolation for genotyping from mouse tail or ear tissue
2-3mm of tail or ear mark is collected into a clean micro-tube or 96-well plate.
Add 75 ul of lysis buffer and seal the tube or plate. If using a heated rack place tubes open for few seconds before closing them again, this will prevent popping up by pressure accumulation.
Incubate plate or tubes at 95oC for 10-60 minutes (15 mins is OK) and cool on ice for 5 minutes.
Add 75 ul of Tris-HCl- UNpH-ed buffer
Mix and centrifuge tubes or plates 5 minutes at 3000 rpm to pellet debris and tissue (which looks nearly intact). Place the supernatant in a new tube –this was not in the original protocol but I found it critical to get reproducible results.
Use 1-2 ul of the supernatant for each PCR reaction. Keep samples frozen when not in use. Thaw each time you need to set up a PCR.
Lysis Buffer:
14 uL conc. NaOH (10N)
14 uL 0.5 M EDTA pH 8.0
dH2O up to 10 ml
Un-pHed Tris-HCl buffer:
40mM TRIS-HCl in dH2O
Reference
BioTechniques Vol 29 No 1 (2000) p52-54
for the isolation of DNA.....in a colony of organisms, we have to get the GENETIC SIGNATURE of the desired organism.
if you are planning to do that by PCR (genotyping) then the above method might work well for you. It is worth testing it as it is very very simple!
Dear Adolfo sir
ok thanks for your comment sir, But i taken in marine microscopic worm so how to possible ur methods,?
We use the method for genotyping mice and the samples are very small (^2 mm2 ear tissue). I guess you can pellet the worms by centrifugation and adjust the volumes to some 20 ul instead of 75 ul of the buffers. End-volume would be e.g. 40 ul and you can use 5-10 ul for PCR.
At the end you need very little DNA, you have PCR for amplifying it!
I worked for DNA extraction techniques, but the first thing to do is to nematodes and sampling should be standardized to use a line.
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