Institute of Cytology and Genetics, Siberian Branch, Russian Academy of Sciences, Novosibirsk, Russia.
University of Haifa, Haifa, Israel.
BMC Genomics. 2018 Feb 9;19(Suppl 3):80. doi: 10.1186/s12864-018-4470-y.
The IWGSC strategy for construction of the reference sequence of the bread wheat genome is based on first obtaining physical maps of the individual chromosomes. Our aim is to develop and use the physical map for analysis of the organization of the short arm of wheat chromosome 5B (5BS) which bears a number of agronomically important genes, including genes conferring resistance to fungal diseases.
A physical map of the 5BS arm (290 Mbp) was constructed using restriction fingerprinting and LTC software for contig assembly of 43,776 BAC clones. The resulting physical map covered ~ 99% of the 5BS chromosome arm (111 scaffolds, N50 = 3.078 Mb). SSR, ISBP and zipper markers were employed for anchoring the BAC clones, and from these 722 novel markers were developed based on previously obtained data from partial sequencing of 5BS. The markers were mapped using a set of Chinese Spring (CS) deletion lines, and F2 and RICL populations from a cross of CS and CS-5B dicoccoides. Three approaches have been used for anchoring BAC contigs on the 5BS chromosome, including clone-by-clone screening of BACs, GenomeZipper analysis, and comparison of BAC-fingerprints with in silico fingerprinting of 5B pseudomolecules of T. dicoccoides. These approaches allowed us to reach a high level of BAC contig anchoring: 96% of 5BS BAC contigs were located on 5BS. An interesting pattern was revealed in the distribution of contigs along the chromosome. Short contigs (200-999 kb) containing markers for the regions interrupted by tandem repeats, were mainly localized to the 5BS subtelomeric block; whereas the distribution of larger 1000-3500 kb contigs along the chromosome better correlated with the distribution of the regions syntenic to rice, Brachypodium, and sorghum, as detected by the Zipper approach.
The high fingerprinting quality, LTC software and large number of BAC clones selected by the informative markers in screening of the 43,776 clones allowed us to significantly increase the BAC scaffold length when compared with the published physical maps for other wheat chromosomes. The genetic and bioinformatics resources developed in this study provide new possibilities for exploring chromosome organization and for breeding applications.
国际小麦基因组测序联盟（IWGSC）构建小麦参考基因组序列的策略是首先获得各个染色体的物理图谱。我们的目标是开发和利用该物理图谱，分析携带多个农艺重要基因（包括抗真菌病基因）的小麦 5B 染色体短臂（5BS）的结构。
利用限制性酶切指纹图谱和 LTC 软件对 43776 个 BAC 克隆进行连续克隆群组装，构建了 5BS 臂（290 Mbp）的物理图谱。该物理图谱覆盖了 5BS 染色体臂的~99%（111 个支架，N50=3.078 Mb）。SSR、ISBP 和拉链标记物被用于 BAC 克隆的锚定，在此基础上，基于 5BS 部分测序获得的先前数据，开发了 722 个新标记物。利用中国春（CS）缺失系、CS 与 CS-5B 二粒小麦的杂交后代 F2 和 RICL 群体，将这些标记物进行了作图。我们采用三种方法将 BAC 连续群锚定在 5BS 染色体上，包括逐个 BAC 筛选、基因组拉链分析和比较 5B 假拟二粒小麦的 BAC 指纹图谱与虚拟指纹图谱。这些方法使我们能够达到较高的 BAC 连续群锚定水平：96%的 5BS BAC 连续群位于 5BS 上。染色体上连续群的分布揭示了一个有趣的模式。包含串联重复区中断区域标记物的短 BAC 连续群（200-999 kb）主要定位于 5BS 端粒区；而较大的 1000-3500 kb BAC 连续群在染色体上的分布与通过 Zipper 方法检测到的与水稻、短柄草和高粱同源的区域分布更好地相关。
与其他小麦染色体公布的物理图谱相比，高指纹图谱质量、LTC 软件和通过信息标记物筛选的 43776 个克隆中大量选择的 BAC 克隆，显著增加了 BAC 支架的长度。本研究开发的遗传和生物信息学资源为探索染色体结构和育种应用提供了新的可能性。
