Singh Nripendra Vikram, Patil Prakash Goudappa, Sowjanya Roopa P, Parashuram Shilpa, Natarajan Purushothaman, Babu Karuppannan Dhinesh, Pal Ram Krishna, Sharma Jyotsana, Reddy Umesh K
ICAR-National Research Centre on Pomegranate (NRCP), Solapur, India.
Gus R. Douglass Institute and Department of Biology, West Virginia State University, West Virginia, WV, United States.
Front Genet. 2021 Jul 28;12:704075. doi: 10.3389/fgene.2021.704075. eCollection 2021.
Here we report on comprehensive chloroplast (cp) genome analysis of 16 pomegranate ( L.) genotypes representing commercial cultivars, ornamental and wild types, through large-scale sequencing and assembling using next-generation sequencing (NGS) technology. Comparative genome analysis revealed that the size of cp genomes varied from 158,593 bp (in wild, "1201" and "1181") to 158,662 bp (cultivar, "Gul-e-Shah Red") among the genotypes, with characteristic quadripartite structures separated by a pair of inverted repeats (IRs). The higher conservation for the total number of coding and non-coding genes (rRNA and tRNA) and their sizes, and IRs (IR-A and IR-B) were observed across all the cp genomes. Interestingly, high variations were observed in sizes of large single copy (LSC, 88,976 to 89,044 bp) and small single copy (SSC, 18,682 to 18,684 bp) regions. Although, the structural organization of newly assembled cp genomes were comparable to that of previously reported cp genomes of pomegranate ("Helow," "Tunisia," and "Bhagawa"), the striking differences were observed with the lines, viz., (NC_0346620) and (NC_031414), which clearly confirmed previous findings. Furthermore, phylogenetic analysis also revealed that members outside the genus were clubbed into a separate clade. The contraction and expansion analysis revealed that the structural variations in IRs, LSC, and SSC have significantly accounted for the evolution of cp genomes of and over the periods. Microsatellite survey across cp genomes resulted in the identification of a total of 233 to 234 SSRs, with majority of them being mono- (A/T or C/G, 164-165 numbers), followed by di- (AT/AT or AG/CT, 54), tri- (6), tetra- (8), and pentanucleotides (1). Furthermore, the comparative structural variant analyses across cp genomes resulted in the identification of many varietal specific SNP/indel markers. In summary, our study has offered a successful development of large-scale cp genomics resources to leverage future genetic, taxonomical, and phylogenetic studies in pomegranate.
在此，我们通过使用下一代测序（NGS）技术进行大规模测序和组装，报告了对16个石榴（L.）基因型（代表商业品种、观赏型和野生型）的叶绿体（cp）基因组的综合分析。比较基因组分析表明，在这些基因型中，cp基因组的大小从158,593 bp（野生型“1201”和“1181”）到158,662 bp（品种“Gul-e-Shah Red”）不等，具有由一对反向重复序列（IRs）分隔的典型四分体结构。在所有cp基因组中，编码和非编码基因（rRNA和tRNA）的总数及其大小以及IRs（IR-A和IR-B）具有较高的保守性。有趣的是，在大单拷贝（LSC，88,976至89,044 bp）和小单拷贝（SSC，18,682至18,684 bp）区域的大小上观察到高度变异。虽然新组装的cp基因组的结构组织与先前报道的石榴cp基因组（“Helow”、“突尼斯”和“Bhagawa”）相当，但与品系（NC_0346620）和（NC_031414）存在显著差异，这清楚地证实了先前的发现。此外，系统发育分析还表明，石榴属之外的成员被归为一个单独的分支。收缩和扩张分析表明，IRs、LSC和SSC的结构变异在很大程度上解释了石榴和在不同时期cp基因组的进化。对cp基因组的微卫星调查共鉴定出233至234个SSR，其中大多数是单核苷酸（A/T或C/G，164 - 165个），其次是二核苷酸（AT/AT或AG/CT，54个）、三核苷酸（6个）、四核苷酸（8个）和五核苷酸（1个）。此外，对cp基因组的比较结构变异分析导致鉴定出许多品种特异性SNP/插入缺失标记。总之，我们的研究成功开发了大规模的cp基因组学资源，以促进石榴未来的遗传、分类和系统发育研究。
