| 摘要: |
| 以桑科榕属植物为研究对象,利用改进的CTAB法提取基因组DNA,并通过单因素多水平梯度试验,筛选了模板、Mg2+、Taq酶、dNTPs和随机引物的浓度及用量,建立了榕属植物RAPD技术分析体系。反应体系总体积为25 μL,各组分浓度为:10×Buffer 2.5μL,Mg2+ 2.5 mmol/L,Taq DNA polymerase 0.06 U/μL,dNTPs 0.2 mmol/L,Primer 0.2 μmol/L．Template DNA 1.2 ng/μL。PCR循环程序为:94 ℃预变性4 min;94 ℃循环变性 1 min,36 ℃退火1 min,72 ℃延伸2 min,40个循环;最后72 ℃延伸8 min。 |
| 关键词: 榕属植物 DNA提取 体系优化 |
| DOI： |
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| Isolation genomeic DNA and optimization of RAPD in Ficus |
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| Abstract: |
| An improved DNA extraction protocol with CTAB was developed for Ficus species.The genomic DNA was employed to establish optimization of RAPD.The RAPD system for Ficus L.(Moraceae) was optimized including program,template,primer,dNTPs,Taq polymerase and Mg2+.Through ladder experiments,the reliable RAPD analysis system is established.The total reaction volume is 25 μL and reaction mixture consist of 2.5 μL 10×Buffe,1.2 ng/μL Template DNA，0.2 mmol/L dNTPs,2.5 mmol/L Mg2+,0.2 μmol/L random Primer and 0.06 U/μL Hotstar Taq DNA polymerase.RAPD program is 4 min at 94 ℃ for predenaturation,then 40 cycles of 1 min at 94 ℃ for denaturation,of 1 min at 36 ℃ for annealing,of 2 min at 72 ℃ for extension,finally extension at 72 ℃ for 8 min. |
| Key words: Ficus DNA extraction RAPD protocol optimization |
