摘要
为探寻出更适宜大麻的ISSR反应体系,用以研究大麻的遗传多样性,利用梯度试验对dNTP、TaqDNA聚合酶和引物的浓度,预变性时间和退火温度5个因素进行优化,从而建立了适合大麻的ISSR-PCR反应体系。结果表明,在25μL体系中,dNTP 200μmol/L,模板1 ng/μL,引物200 nmol/L,聚合酶1 U(2.5 pmol/μL);预变性3 min(94℃),退火温度48℃,为大麻最佳反应体系;研究利用优化后的ISSR反应体系,在30条引物中,筛选出8条适合大麻的ISSR引物,体系具有较好的稳定性和重复性。
The five factors affected ISSR reaction, including dNTP, TaqDNA polymerse, concentrations of primer, predegeneration time, annealing temperature etc., were tested to optimize ISSR amplification system by single factor experiment method for finding genetic diversity of hemp. The results showed that： the optimum reaction system in 25 μL volume was dNTP 200 μmol/L, template 1 ng/μ, primer 200 nmol/L, Taq DNA polymerase 1 U （2.5 pmol/μL）, predegeneration time 3 min （94℃）, annealing temperature 48℃. Moreover, 8 primers in 30 primers were screened out through the above stable reaction system. The repeatable and stable system was completely suitable to analyze the hemp genetic information.
出处
《中国农学通报》
CSCD
2014年第12期105-109,共5页
Chinese Agricultural Science Bulletin
基金
黑龙江省自然科学基金项目"大麻种质资源的细胞遗传学研究与ISSR分析"(QC2012C115)
哈尔滨市青年科技创新人才项目"大麻种质资源的遗传多样性研究"(2013RFQYJ014)
