基于生物信息学的睡莲SSR位点特征分析

SSR Loci Characteristic Analysis of Water Lily Based on Bio-informatics Methodology

【目的】分析蓝星睡莲和小花睡莲叶片转录组测序产生的Unigene序列及蓝星睡莲全基因组序列的简单重复序列(SSR)位点特征,为开展睡莲属植物种质资源鉴定、遗传多样性分析及遗传连锁图谱构建等提供基础数据。【方法】利用前期研究获得的蓝星睡莲和小花睡莲叶片转录组数据及已公开发表的蓝星睡莲基因组,以MISA进行SSR位点搜索,并统计分析其全基因组序列的SSR位点出现频率、基元序列长度和基元类型等。【结果】在蓝星睡莲和小花睡莲叶片转录组的114762个Unigenes序列中搜索到38998个SSR位点,SSR位点出现频率为33.98%,其中完整型SSR位点30124个;SSR基元序列总长度为465550 bp,总平均为21.65 bp。在蓝星睡莲基因组中搜索到249029个SSR位点,平均分布频率为609.0个/Mb,其中完整型SSR位点163265个;SSR基元序列总长度为2775181 bp,总平均为27.25 bp,占基因组大小的0.68%。在转录组和基因组中,SSR位点均以二核苷酸和单核苷酸重复类型为主,分别占SSR总数的48.87%和41.03%、51.70%和43.37%;在二核苷酸重复基元中,AG/TC和AT/TA型的占比较高,且远高于其他类型重复基元;SSR基元中各类型重复以5~11次为主,其中重复10次的最多;基于荧光毛细血管电泳法从合成的144对SSR引物中筛选出12对扩增性好且多态性高的SSR引物。【结论】蓝星睡莲和小花睡莲叶片转录组及蓝星睡莲基因组SSR中的低级基元类型较丰富,具有开发为高多态性SSR引物的潜力;筛选出12对扩增性好且多态性高的SSR引物可用于开展睡莲属植物种质资源鉴定、遗传多样性分析及遗传连锁图谱构建等研究。

【Objective】The Unigene produced by leaf transcriptome of Nymphaea colorata and N.micrantha and the whole genome sequence of N.colorata were analyzed according to SSR loci characteristics.It provided basic data for the germplasm resource identification,genetic diversity analysis and genetic linkage map construction of water lily plants.【Method】Based on the leaf transcriptome data of N.colorata and N.micrantha which were acquired previously in the laboratory and the published genome of N.colorata,MISA software was used to search SSR loci and analyze the frequency,length and primitive types of SSR.【Result】38998 SSR loci were found among the 114762 Unigenes sequences obtained from the transcriptome of N.colorata and N.micrantha.The SSR site frequency was 33.98%.Among them,30124 SSR loci belong to complete types.The total length of the SSR motif sequence was 465550 bp,and had a total average of 21.65 bp.249029 SSR sites were searched in the N.colorata genome,with an average distribution frequency of 609.0/Mb,including 163265 intact SSR sites;the total length of SSR motif sequence was 2775181 bp,and 27.25 bp,accounted for 0.68%of N.colorata genome size.In the transcriptome and genome,the SSR loci were mainly dinucleotide and single nucleotide repeats,which respectively accounted for 48.87%,41.03%and 51.70%,43.37%of the total SSR.Among dinucleotide repeat motifs,AG/TC and AT/TA types were higher and both much higher than other types of repeat motifs.The number of repetitions of each type in the SSR primitive was mainly 5 to 11 times,of which 10 times were the most.Twelve pairs of SSR primers with good amplification and high polymorphism were selected based on fluorescence capillary electrophoresis from synthetic 144 pairs of SSR primers.【Conclusion】Low-level primitive types are abundant in the SSR of N.colorata and N.micrantha transcriptome and genome,which has great potential for developing and screening SSR primers with high polymorphism.The selected 12 pairs with good amplifications and high polymorphism of SSR primers can be used to study the identification of germplasm resources,genetic diversity analysis and genetic construction of plant chain map.