Characterization of atpA and orf220 genes distinctively present in a cytoplasmic male-sterile line of tuber mustard
Characterization of atpA and orf220 genes distinctively present in a cytoplasmic male-sterile line of tuber mustard
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块茎芥菜细胞质雄性不育系中独特存在的 atpA 和 orf220 基因的表征
DOI：
10.1080/14620316.2003.11511706
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发表时间：
2003-01
影响因子：
1.9
通讯作者：
Yutaka Hirata等
中科院分区：
文献类型：
--
作者：
张明方;陈利萍;陈竹君;杨景华;Yutaka Hirata等
摘要 在本研究中，编码 F1-ATP 合酶 a 亚基的 atpA（登录号：AY211265、AY211266）基因以及可能编码具有推导的 220 个氨基酸的疏水性膜相关多肽的开放阅读框 orf220（登录号：AY208898）被发现独特地存在于细胞质雄性不育（cms）中。块茎芥菜线。通过使用线粒体特异性引物优先克隆它们并进行部分表征。从 cms 及其可育维持系 (m) 系克隆的 atpA 基因经测序，片段大小分别为 1,454 和 1,073 bp。它们在核苷酸成分方面具有约70%的高度相似性。然而，这两个基因的比对表明，与 m 系相比，cms 系的 atpA 基因在 5' 侧翼插入了 383 bp 大小的片段，横跨第 141 到 523 个碱基位点。此外，还发现开放阅读框orf220仅存在于芥菜的cms品系中。推测其可能编码疏水性N端膜相关多肽，推测分子量为26 kD。 Southern杂交进一步表明orf220仅存在于cms系中，而在可育m系中不存在，未检测到杂交信号。尽管 atpA 基因在 cms 或 m 系中被印迹，但当分别用 Hind III 和 Xba I 消化的 DNA 印迹时，它们具有不同的片段大小。 atpA 和 orf220 基因在线粒体基因组中被认为是低表达的。此外，系统发育分析表明，orf220基因与甘蓝nap cms中的orf222的亲缘关系比甘蓝中orf224和拟南芥中orf240的亲缘关系更密切。这两个基因参与芥菜 cms 表达的直接证据需要进一步研究。
Summary In the present investigation, the atpA (Accession Number: AY211265, AY211266) gene, encoding the a subunit of F1-ATP synthase, as well as the open reading frame orf220 (Accession Number: AY208898), presumably encoding a hydrophobic membrane-associated polypeptide with deduced 220 amino acids, were found to be distinctively present in a cytoplasmic male-sterile (cms) line of tuber mustard. They were preferentially cloned by using mitochondria-specific primers and were partially characterized. The atpA gene cloned from cms and its fertile maintainer (m) lines were sequenced to be 1,454 and 1,073 bp-sized in fragments, respectively. They had a high similarity of approximately 70% in terms of nucleotide constituents. However, alignment of these two genes suggested that the atpA gene from the cms line had an insertion of 383 bp-sized fragment in 5’-flank spanning from 141th to 523th base site, compared with that of its m line. Besides, the open reading frame, orf220, was found to be exclusively present in the cms line of tuber mustard. It was deduced to possibly encode a hydrophobic N-terminus membrane-associated polypeptide with a deduced molecular weight of 26 kD. By Southern hybridization, it was further shown that orf220 existed only in the cms line and was absent from the fertile m line with no hybridization signal detected. Although the atpA gene was blotted in either cms or m line, they were of different fragment sizes when blotted with DNA digested with Hind III and Xba I, respectively. Both atpA and orf220 genes were supposed to be low coped in mitochondria genomes. In addition, by phylogenetic analysis, it was manifest that the orf220 gene had a closer genetic relationship with orf222 in Brassica nap cms than both of orf224 in Brassica pol cms and orf240 in Arabidopsis thaliana cms. The direct evidence of the involvement of these two genes in the expression of cms of tuber mustard needs further investigation.