文献类型：期刊文献

中文题名：基于毛细管电泳的柳树AFLP分子标记研究

英文题名：AFLP molecular marker of Salix spp.based on apillary electrophoresis

作者：吕金辉[1] 胡建军[1] 卢孟柱[1]

第一作者：吕金辉

机构：[1]林木遗传育种国家重点实验室 中国林业科学研究院林业研究所

年份：2012

卷号：34

期号：1

起止页码：51-57

中文期刊名：北京林业大学学报

外文期刊名：Journal of Beijing Forestry University

收录：CSTPCD;;北大核心:【北大核心2011】；CSCD:【CSCD2011_2012】；

基金：“863”国家高技术研究发展计划项目(2011AA100201);转基因生物新品种培育科技重大专项(2009ZX08011-027B)

语种：中文

中文关键词：柳树;AFLP;毛细管电泳

外文关键词：Salix spp.; AFLP; capillary electrophoresis

分类号：S718.46

摘要：为构建柳树遗传图谱、进行分子育种等奠定基础,以柳树为材料,基于毛细管电泳技术体系建立并优化了AFLP分子标记技术,简化了AFLP分析流程。首先提取高质量的柳树基因组DNA,对基因组进行酶切与接头连接、预扩增和选择性扩增,最后通过毛细管电泳分析各因素的影响。基因组DNA提取采用改进的CTAB法,酶切模板DNA用量450ng,EcoRⅠ酶切2h,MseⅠ酶切2h,接头过夜连接,选择性扩增时dNTP浓度0.3mmol/L,Mg2+浓度1.5mmol/L,引物浓度0.125μmol/L,DNA聚合酶浓度0.025U/μL,预扩增产物最适稀释倍数20倍。经过重复实验,证明建立的AFLP-毛细管反应体系适用于柳树AFLP分析。
In this paper,we established and optimized the AFLP molecular marker technology of Salix spp.based on capillary electrophoresis as well as simplify analysis flow of AFLP in order to construct Salix spp.linkage mapping and the implement of molecular breeding.First,the high-quality genomic DNA was extracted for AFLP analysis.Then the genomic DNA was completely digested by restriction enzyme and ligased with oligonucleotide adapters successfully.We got the products via pre-amplification and selective amplification.At last,the selective amplification products were analyzed by capillary electrophoresis.Results showed that improved CTAB method was selected to extract genomic DNA,450 ng of genomic DNA was used in digestion with EcoRⅠ and MseⅠ for 2 hours respectively,and ligased overnight.In selective amplification,0.3 mmol/L dNTP,1.5 mmol/L Mg2＋,0.125 μmol/L primers,0.025 U/μL polymerase and 20 times dilution of pre-amplification products were desirable.Through many repeats,this system has been proved to be feasible in Salix spp.AFLP-capillary electrophoresis analysis.