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对影响中国水仙“金盏”品种的REMAP-PCR扩增反应的各个参数进行优化,建立适合中国水仙的REMAP反应体系20μL,包括如下成分:模板DNA 40 ng、dNTPs 0.2 mmol.L-1、引物0.45μmol.L-1、Taq酶1U和10×Buffer(含Mg2+)2μL。扩增程序为反应程序:94℃预变性4 min;94℃变性40 s,55℃退火40 s;72℃延伸90 s,共35个循环;最后72℃延伸10 min,4℃保存。该反应体系标记点位清晰、稳定、重复性好,适宜中国水仙的REMAP分析,为应用REMAP技术鉴定中国水仙种质资源、分子标记辅助选择育种及其遗传多样性研究奠定了基础。
The parameters of REMAP-PCR amplification reaction affecting Chinese narcissus cultivar “Calendula” were optimized, and 20μL of REMAP reaction system for Chinese narcissus was established, including the following components: template DNA 40 ng, dNTPs 0.2 mmol.L-1 , Primer 0.45μmol.L-1, Taq enzyme 1U and 10 × Buffer (containing Mg2 +) 2μL. The amplification procedure was as follows: pre-denaturation at 94 ° C for 4 min, denaturation at 94 ° C for 40 s, annealing at 55 ° C for 40 s and extension at 72 ° C for 90 s for a total of 35 cycles and final extension at 72 ° C for 10 min and preservation at 4 ° C. The REMAP analysis of Chinese narcissus is a clear, stable and reproducible marker for the reaction system, which lays the foundation for the identification of Chinese Narcissus germplasm resources, molecular marker-assisted selection breeding and genetic diversity using REMAP.