适用于微卫星标记的湿地松、加勒比松DNA快速提取法

Study of Rapid DNA Extraction Method Suitable for SSR Analysis in Slash Pine and Caribbean Pine

以湿地松(Pinuselliottii,PEE),包括古巴加勒比松(P.caribaea var.caribaea,PCC)、洪都拉斯加勒比松(P.caribaea var.hondurensis,PCH)、巴哈马加勒比松(P.caribaeavar.bahmaensis,PCB)3个变种在内的加勒比松,侧枝顶芽为试验材料,使用RETSCHMM400混合球磨仪破碎植物组织,然后利用改良CTAB法提取基因组DNA,完成48个样品仅用1h,1个工作日可提取300个样品以上,DNA分子量大于20kb,0.3g样品可提取DNA20-60μg,能够满足几百次PCR反应;利用所提DNA进行SSR分析,条带清晰,多态性好,说明该方法提取的DNA完全可以满足SSR反应的需要。本研究为SSR标记用于湿地松、加勒比松分子标记辅助选择育种提供了经济、高效、可靠的DNA提取方法。

One improved CTAB method to extracted genomic DNA from terminal buds on lateral branch of slash pine and Caribbean pine was provided. According to the improved method and the use of RETSCH MM400 fragmented plant tissue for extracted of DNA, the efficiency was raised compared to the normal process. In the expertment,48 samples could be treated one time and the process could be finished in one hour,and DNA of 300 samples could be obtained in one workday. It was estimated that molecular weight of the extracted DNA is larger than 20 kb. And 20 -60 g DNA could be extracted from 0. 3 g terminal bud,which would be enough for hundreds of times of PCR reaction. The extracted DNA was using in SSR analysis,the banding patterns was clear and with good polymorphism, and the quality of DNA extracted using this method was suitable for SSR analysis. This study provide an economic,efficient, and reliable DNA extraction method for SSR markers use for molecular marker-assisted selection breeding of Slash pine,Caribbean pine.