1. The purpose of the experiment
2. Experimental principle
DNA is genetic material, and the extraction of plant DNA is the basis of plant genetic engineering. It is usually required that the extracted DNA has ideal purity, sufficient integrity, no breakage and degradation, and minimal use of toxic chemicals in the extraction process. In order to meet the above requirements, many methods of extracting plant DNA have been reported. In summary, there are mainly CTAB method, SDS method and alkaline extraction method. DNA molecule is the basic material of molecular biology research. Different extraction methods can be used to obtain DNA of varying quantity and quality according to different experimental purposes. Therefore, the ratio of reagents in the extraction solution is also different: the extraction of plant DNA often uses the CTAB method.
The formula is as follows:
1. Before preparing the extract, prepare the mother liquor.
Mother liquor (sterilized): 5M NaCl (58.4g add ddH 2 O to make 200ml solution)
0.5M Na 2 -EDTA (37.22g+ddH 2 O to 200ml, adjust the pH to 8.0 with NaOH)
1M Tris-HCl (12.11g Tris-HCl+ ddH 2 O to 100ml, adjust the pH to 8.0 with concentrated hydrochloric acid)
2. Extraction solution formula
100ml
Add: CTAB powder 2g
1M Tris(pH8.0) 10ml
0.5M EDTA(pH8.0) 4ml
5M NaCl 28ml
PVP 1g
β-mercaptoethanol 1ml
Add double distilled water to make the volume to 100ml
3. Extraction solution
Measure phenol, chloroform, and isoamyl alcohol and mix them in a volume ratio of 25:24:1, then measure chloroform and isoamyl alcohol and mix them in a volume ratio of 24:1 to prepare two extracts, each with 100ml
4 0.1xTE
1ml Tris, 20µl EDTA with H 2 O, adjust the pH to 8.0 with HCl, fix the solution, and sterilize
Three, experimental equipment
Beaker, measuring cylinder, volumetric flask, glass rod, straw, pipette, centrifuge tube, refrigerator, autoclave, electronic balance, bottle washing
Four, experimental reagents
Distilled water, CTAB, NaCl, EDTA, Tris-HCl, β-mercaptoethanol, 0.1xTE, RNase enzyme, phenol, chloroform, isoamyl alcohol, absolute ethanol
In summary, it can be seen that manual extraction of DNA is a very complicated and tedious process. If you use DNA extractor, RNA extractor, DNA extraction kit, and RNA extraction kit, you can complete the experiment more efficiently.
