大花序桉基因组SSR的分布特征及序列分析

【目的】分析大花序桉基因组SSR的分布特征及序列分析，为大规模开发大花序桉分子标记辅助育种的SSR标记提供理论依据。【方法】通过高通量测序获得大花序桉基因组序列信息，经严格过滤、拼接和组装后获得scaffolds，利用MISA网站对Scaffolds进行SSR位点检索及分析，并利用Primer 5.0对具有较大多态性开发潜力（二基元重复次数≥10、三基元重复次数≥7）的SSR标记进行引物设计，随机挑选48对基因组SSR引物进行有效性检测及有效扩增引物筛选。【结果】大花序桉基因组共含有177750个SSR位点，包括171530个SSR独立位点和6220个复合型SSR位点；SSR发生频率为17.86%，分布密度为1/3.13 kb。大花序桉基因组SSR基元类型较丰富，以单核苷酸重复基元类型数量最多，占基因组总SSR数量的69.33%，其次为二、三核苷酸重复基元类型，分别占基因组SSR数量的21.01%和7.58%，四、五、六核苷酸的重复基元类型所占比例均相对较低，三者的比例含量总和为2.08%。SSR基元类型以AG/CT占绝对优势（16812个），其次为AT/AT（8193个）和AAG/CTT（4404个）。从48对SSR标记引物中筛选出31对引物能有效扩增出清晰、明亮条带且大小与预期相符，其中有14对在6个大花序桉样本中均呈现多态性，多态百分率为29.17%。【结论】大花序桉基因组SSR位点发生频率高，基元类型较丰富，SSR多态性标记开发潜力大。该研究结果为进一步大花序桉SSR引物开发和筛选，以及开展大花序桉及其他桉树遗传多样性分析和遗传图谱构建提供依据。

Characteristics and analysis of simple sequence repeats(SSR)in Eucalyptus cloeziana genome

【Objective】The distribution patterns and sequence characterstics of simple sequence repeat(SSR) in Eucalyptus cloeziana genome were analyzed in order to lay the foundation for developing SSR markers in molecular marker-assisted breeding of E. cloeziana.【Method】The genome sequence information of E. cloeziana was obtained based on high-throughput sequencing. Scaffold sequences were identified after rigorous filtration, splicing and assembly. The SSR loci from Scaffolds were searched and analyzed by using MicroSAtellite(MISA). Primer 5.0 software was used to design the SSR sequences with high potential for polymorphism development(the repetitions of dinucleotide ≥ 10, the repetitions of trinucleotides ≥ 7). Forty-eight genomic SSR primer pairs were randomly selected for validity detection and screening of primers successfully amplicated.【Result】The genome of E. cloeziana contained 177750 SSR loci, including 171530 single SSRs and 6220 compound-loci SSRs. The SSR occurrence frequency was 17.86% and the distribution density was 1/3.13 kb. There were abundant SSR motifs in E. cloeziana genome, and among them, the mononucleotide repeats were the most abundant types, accounting for 69.33% of the total genomic SSRs, followed by dinucleotide and trinucleotide repeats, accounting for 21.01% and 7.58%, respectively. The proportions of tetra -, penta-and hexa-nucleotide repeats were relatively low, and the total proportion of the three nucleotides was 2.08%. For the SSR motifs, AG/CT were the dominant primitive types(16812), followed by AT/AT(8193), AAG/CTT(4404). A total of 48 SSR marker pairs were randomly selected for initial screening test. Thirty-one primers pairs could effectively amplify clear and bright bands matching the expected size, and 14 of them showed polymorphism across six samples of E. cloeziana. The percentage of polymorphic loci was 29.17%.【Conclusion】The SSR loci in the genome of E. cloeziana occur in high frequency and abundant motifs, and exhibit great potential for developing polymorphic SSR markers. Therefore, the results provide a basis for further SSR development and screening, genetic diversity analysis and genetic mapping of E. cloeziana and other Eucalyptus trees.