Determination of the entire structure of wheat chloroplast genome

叶绿体基因组全序列仅在水稻和玉米两种禾本科植物中报道。在本项目中，作为第三个成功测定小麦叶绿体基因组全序列的案例。我们使用一个普通小麦（小麦）品种。中国春作为材料，因为这个品种已被广泛用于世界各地的遗传调查。此外，我们还构建了小麦叶绿体DNA重叠群克隆库。此外，PCR引物设计，以扩增所有的，但两个小麦叶绿体基因的编码区，使用它成功地扩增其编码区。主要结果如下。(1)小麦叶绿体基因组全序列：小麦叶绿体基因组大小为134，540 bp，比玉米（140，387 bp）小4.2%，与水稻（134，525 bp）基本一致。大小单拷贝区（LSC）、小单拷贝区（SSC）和反向重复区（IR）的大小分别为80，347、12，789和20，702 bp。该基因组包含109个基因，其中4个（重复）编码4种rRNA，30个编码tRNA（对应于20个氨基酸），70个编码蛋白质亚基，5个功能不明。(2)重叠群克隆库：构建了一个由55个重叠群克隆组成的叶绿体DNA库，这些克隆覆盖了LSC、SSC和IR的整个区域。克隆的平均大小为2，246 bp。(3)小麦叶绿体基因的PCR扩增引物组合：为107个叶绿体基因设计了一对PCR引物。使用这些，少量的扩增DNA制备的编码区的所有，但两个小麦叶绿体基因。

Complete nucleotide sequence of chloroplast genome has been reported only in two Gramineae species, rice and corn. In this project, we succeeded determination of the complete sequence of wheat chloroplast genome as the third case. We used a common wheat (Triticum aestivum) cv. Chinese Spring as the material, because this cultivar has been used most extensively worldwide in genetic investigations. In addition, we constructed a contig clone bank of wheat chloroplast DNA. Also, PCR-primer sets were designed to amplify coding regions of all but two wheat chloroplast genes, using which their coding regions were successfully amplified. Main results are given below.(1) Complete sequence of wheat chloroplast genome : Size of wheat chloroplast genome was 134,540 bp, that was 4.2% smaller than that of corn (140,387 bp) but almost the same as that of rice (134,525 bp). Sizes of three regions, large and small single copy regions (LSC and SSC) and inverted repeat (IR), were 80,347, 12,789 and 20,702 bp, respectively. This genome contained 109 genes, 4 (in duplication) coding for 4 rRNA species, 30 for tRNAs (corresponding to 20 amino acids), 70 for protein subunits and 5 of unidentified functions.(2) Contig clone bank : A chloroplast DNA bank consisting of 55 contig clones (some are partially overlapping) that cover entire regions of LSC, SSC and IR was constructed. Average size of the clones was 2,246 bp.(3) PCR primer sets for amplification of wheat chloroplast genes : A pair of PCR primers for each of 107 chloroplast genes were designed. Using these, small amounts of amplified DNAs were prepared for the coding regions of all but two wheat chloroplast genes.