甘蔗单花粉全基因组扩增(WGA)与SCoT分子标记研究  

Whole Genome Amplification of Single Pollen Grains from a Sugarcane Cultivar and Analysis of the Genetic Relatedness based on SCoT Markers

作　　者：陈平华[1] 陈如凯[1] 许莉萍[1] 王恒波[1] 陈利平[1] 林冰[1] 施桂姣[1] 张卓[1] 高三基[1] 郭晋隆[1] 潘永保[1] 

机构地区：[1]福建农林大学农业部甘蔗遗传改良重点开放实验室,福建福州350002

出　　处：《热带作物学报》2011年第11期2069-2075,共7页Chinese Journal of Tropical Crops

基　　金：国家基金项目(No.31070330);国家948项目(No.2010-S19);福建省科技计划重点项目(No.2007I0036)

摘　　要：从一个完整的甘蔗花粉囊中分离出单个花粉粒．利用特制取样器进行收集，采用碱裂解法释放单花粉DNA．裂解液可以直接作为WGA模板进行全基因组扩增．制备出单个花粉粒大量DNA。通过5SrRNA—ITS鉴定WGA产物真实性．并以目标起始密码子多态性标记对真实的WGA产物进行分析，显示出丰富的多态性。利用NTSYSpc软件分析花粉粒间遗传距离，并进行聚类分析，显示单花粉间遗传距离变幅为0．0888-0．3654，在相似系数0．187处．受试花粉粒中的36个明显分为2组．占总数的69％。说明减数分裂染色体重组过程中，有一定比例的基因位点为多数花粉粒所共有。根据SCoT标记聚类图可将所有供试花粉分为4大类．不同类花粉粒的分布具有一定的规律性．Single pollen grains were isolated from an intact anther of a sugarcane cultivar and collected using a pair of special forceps. The single pollen grains were lysed in an alkali/detergent solution respectively. The resulting solution was used as the template for Whole Genome Amplification. Genomic DNA of individual pollen grains was amplified successfully and WGA products of single pollen grains were tested by 5S rRNA-ITS primers to select those with high fidelity to the donor plant. The WGA products selected out were analyzed with SCoT markers and produced a mount of DNA polymorphism. Genetic similarity calculation was performed using the software NTSYSpc 2.1. The similarity coefficient values ranged from 0.0888N0.3654 and 69 percent of total pollen grains were clustered into two groups at the value of 0.187.The results indicated a high scale of pollen grains shared same gene loci during the meiotic ,segregation stage. The 52 single pollen grain samples were clustered into four groups based on the homology tree. A probability distribution of the pollen grains was calculated based on the number of pollen grains in the four groups.

关 键 词：甘蔗 单花粉 全基因组扩增 SCoT分子标记 遗传分布 

分 类 号：S566.1[农业科学—作物学]