I am doing a DNA extraction of blood of Mergus octosetaceus with phenol-chloroform, and when I put the TrisNH4Cl (5M) 1000ul, it makes the solution homogeneous, so even after I centrifuge the sample, it doesn't form pellets so I can continue the protocol and add the saline solution. It's important to highlight that the blood sample was being stored with EDTA, so I'm thinking that the Tris is not the best lysis buffer. Has anyone done a dna extraction of blood of birds and can help me??
Too much salt in the mixture can cause phase inversion if using only phenol rather than phenol chloroform but homogeneity is unexpected. Check that your phenol is straw coloured not pink or brown (too old and poor quality) and make sure that your phenol is pH adjusted to about 8 with your tris buffer. You cannot use pH papers for this but 1ml of phenol in 9ml of 45% methanol should work with a pH meter
I haven't worked with bird blood, but I remember reading in the Qiagen DNeasy kit that since bird red cells still have nuclei that it's easy to have too much starting material.
Not sure if that helps narrow down the list of possibilities, but thought I'd toss it out there just in case the original protocol was for mammal samples (e.g. mouse).
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