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Optimization of Polymerase Chain Reaction Conditions of Denaturing Gradient Gel Electrophoresis for North Black Soil Microbes
| Content Provider | Semantic Scholar |
|---|---|
| Author | Wang, Siyu Wu, Feng Liu, Shou |
| Copyright Year | 2009 |
| Abstract | This paper studies the effects of concentrations of Mg and dNTP, the annealing temperature, extension and cycling times in PCR of north black soil microbes using an orthogonal experiment. Results showed that the feasible PCR reaction system for soil microbes should be carried in a volume of 50 l, composed of 50 ng soil microbial DNA template, 15 pm primer, 5 u Pfu enzyme, 0.2 mmol.L Mg2SO4, and 0.3 mmol.L dNTPs. The PCR reaction procedures were set up under two conditions. At first, the soil microbial DNA was denatured at 94°C for 5 min followed by 20 cycles of each 94°C for 1 min, 65-55°C for 1 min (descending at 0.5°C per cycle), and finally 72°C for 1 min. The second reaction conditions also included 20 cycles, each of 94°C for 1 min, 56°C for 1 min, and 72°C for 1 min, and a final extension temperature of 72°C for 7 min. The work was repeated several times on the same model using the same primers and PCR conditions to acquire clear and pure bands. _ |
| File Format | PDF HTM / HTML |
| Alternate Webpage(s) | http://www.globalsciencebooks.info/Online/GSBOnline/images/0706/DSDP_1(1)/DSDP_1(1)54-57o.pdf |
| Language | English |
| Access Restriction | Open |
| Content Type | Text |
| Resource Type | Article |
