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作 者 ：马喜军1,2,龚桂芝2,彭祝春2,韩学智1,洪棋斌2*

期 刊 ：西北植物学报 2012年 6期 页码：1112~1117

关键词：柑桔；遗传图谱；分子标记；SSR；

Keywords:citrus, genetic linkage map, molecular marker, SSR,

摘 要 ：为了系统性地开发和拓展柑桔SSR标记，通过对公布的柑桔BAC文库末端序列(BAC End sequence，BES)进行SSR分析，选择1500个SSR位点设计合成并检测323对引物。结果表明：（1）从总长度为28.1 Mb的46 339条序列中共检测出22 403个SSR位点，约每2条序列就会出现一个SSR位点，发生频率为48%，相当于平均1.25 kb的序列中就会出现1个SSR，频率约为柑桔EST的2倍，且不同核心重复序列的SSR发生特点与EST也不同。（2）所合成的323对引物中，有效扩增316对，扩增率约98%，173对表现多态性，总多态性比率约55%，多态性引物中单核苷酸重复类型15对，双核苷酸重复类型100对，三核苷酸及以上重复类型58对，表明柑桔BES中具有较为丰富的多态性SSR标记。（3）结合已发表的遗传作图数据，对总计349个多态性位点进行遗传连锁分析，获得的新遗传连锁图谱共含有9个连锁群、334个SSR标记、总长844.2 cM、平均图距2.53 cM，延长和加密了先前的图谱。该研究开发的新SSR标记为开展柑桔遗传鉴定分析和遗传图谱构建提供了新的标记来源，加密的遗传图谱为柑桔的基因定位、图位克隆和标记辅助育种等奠定了基础，SSR分析结果也为其他物种SSR标记的开发提供了参考。

Abstract:Simple Sequence Repeats (SSR) were surveyed in citrus BAC End sequence (BES) for the development of new SSR markers to extend and saturate existing genetic map.22 403 SSRs with 1～6 bp motif were identified in 46 339 citrus BESs retrieved from NCBI.The estimated frequency of SSRs was approximately 1/1.25 kb,nearly twice the frequency found in Citrus Expressed Sequence Tags.323 primers were selected in the present mapping study,of which 40 were mono nucleotide repeats,184 di nucleotide repeats,99 tri nucleotide or above repeats.Polymorphism tests showed that 316 primer pairs (98%) could be amplified successfully and 173 pairs (55%) were polymorphic.Among the polymorphic primers,15 pairs were of mono nucleotide repeats,100 pairs of di nucleotide repeats,58 pairs of tri nucleotide or above repeats.A new linkage map was produced by combining the segregating data of new markers and markers in the previous map.When compared with the published map,the new map integrated 334 SSR markers,9 linkage groups and covered 844.2 cM of citrus genome with an average genetic distance standing at 2.53 cM.The results demonstrated that citrus BES was good for SSR marker development and the developed markers were useful in extending and saturating the citrus genetic maps.So this will provide a reference for development of other species SSR markers and lay a good foundation for citrus gene mapping,map based cloning and marker assisted breeding.

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