Method for extracting metagenome DNA from endosymbiotic bacterium mycetocyte of bemisia tabaci
A technology of metagenomics and symbiotic bacteria, which is applied in the field of extracting metagenomic DNA, can solve the problems of difficult extraction, and achieve the effect of easy implementation, economical cost saving and simple implementation
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Embodiment 1
[0054] step 1:
[0055] The endosymbiont bacteria cell extract containing Bemisia tabaci was used denaturing solution (denaturing solution consisted of: 4M guanidine isothiocyanate, pH 7.0 10mM Tris-HCl buffer, 1mM EDTA, volume final concentration of 0.5 % 2-mercaptoethanol) treatment, the wet weight-volume ratio of the whitefly endosymbiont bacterium cell extract to the denaturation solution is 1g: 1mL to obtain a mixed solution;
[0056] Step 2:
[0057] Add nucleic acid extraction buffer (nucleic acid extraction buffer consists of: 10mM sodium phosphate buffer with pH 7.5, 10mM Tris-HCl buffer with pH 7.5, 10mM Tris-HCl buffer with pH 8.0) in the sample processed in step 1. EDTA, the NaCl of 1.5M, the CTAB that mass final concentration is 1%, the sucrose that mass final concentration is 10%) and enzyme (enzyme is proteinase K and lysozyme use simultaneously, wherein the amount that adds proteinase K is after adding proteinase K Final concentration reaches 30 μ g / mL, and t...
Embodiment 2
[0067] step 1:
[0068] The endosymbiont bacterial cell extract containing Bemisia tabaci was denatured solution (denaturation solution is composed of: 6M guanidine isothiocyanate, pH is 7.5 10mM Tris-HCl buffer solution, 1mM EDTA, volume final concentration is 0.5 % 2-mercaptoethanol) treatment, the wet weight-volume ratio of the whitefly endosymbiont bacterium cell extract to the denaturation solution is 1g: 2mL to obtain a mixed solution;
[0069] Step 2:
[0070] Add nucleic acid extraction buffer (nucleic acid extraction buffer consists of: 10mM sodium phosphate buffer with pH 7.5, 10mM Tris-HCl buffer with pH 7.5, 10mM Tris-HCl buffer with pH 8.0) in the sample processed in step 1. EDTA, the NaCl of 1M, the CTAB that mass final concentration is 1%, the sucrose that mass final concentration is 5%) and enzyme (enzyme is that proteinase K and lysozyme are used simultaneously, and wherein the amount of adding proteinase K is that after adding proteinase K, its final Concen...
Embodiment 3
[0080] step 1:
[0081] The endosymbiont bacteria cell extract containing Bemisia tabaci was used denaturing solution (denaturing solution consisted of: 5M guanidine isothiocyanate, pH 8.0 10mM Tris-HCl buffer, 1mM EDTA, the volume final concentration was 0.5 % 2-mercaptoethanol) treatment, the wet weight-volume ratio of the whitefly endosymbiont bacterium cell extract to the denaturation solution is 1g: 1.5mL to obtain a mixed solution;
[0082] Step 2:
[0083] Add nucleic acid extraction buffer (nucleic acid extraction buffer consists of: 10mM sodium phosphate buffer with pH 7.5, 10mM Tris-HCl buffer with pH 7.5, 10mM Tris-HCl buffer with pH 8.0) in the sample processed in step 1. EDTA, the NaCl of 2M, the CTAB that mass final concentration is 1%, the sucrose that mass final concentration is 8%) and enzyme (enzyme is that proteinase K and lysozyme are used simultaneously, and wherein the amount of adding proteinase K is its final after adding proteinase K Concentration re...
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