关键词: 小麦, 细胞分裂素氧化/脱氢酶, 基因克隆, 染色体定位, 遗传作图

Abstract:

Phytohormone cytokinin (CK) has been shown to influence various aspects of plant growth and developmental events including shoot and root branching, leaf development, the release of buds from apical dominance, the delay of senescence, the promotion of seed germination, and chloroplast formation. Cytokinin oxidase/dehydrogenases (CKX) catalyze the irreversible degradation of the cytokinins isopentenyladenine, zeatin, and their ribosides in a single enzymatic step by cleavage of the unsaturated N6-isoprenoid side chains. This enzyme probably plays the principal role in controlling CK levels in plant tissues. So it is conceivable that genetic manipulation of CKX improves agricultural traits, such as yield attributes or adaptation to environmental stress in crops. In the present work, a novel gene that encodes cytokinin oxidase/dehydrogenase (TaCKX2) was cloned from hexaploid wheat (Triticum aestivum) through the homologous cloning method, and a SSR marker for TaCKX2 was developed according to 3’-UTR sequence of TaCKX2. Sequence analysis showed that the complete ORF of TaCKX2 was 1 554 bp, and encodes a putative protein composed of 517 amino acids. Sequence alignment proved TaCKX2 shared the highest homology with rice OsCKX11 (85.3% cDNA sequence identity) instead of OsCKX6 as previously reported. A series of ‘Chinese Spring’ nulli-tetrasomic stocks were employed to ascertain the chromosomal location of TaCKX2, the result showed that TaCKX2 was on wheat chromosome 7B and 7D. Using a population of 199 recombinant inbred lines from the cross Yanzhan1×Neixiang188, TaCKX2 was mapped between SSR markers Wmc316 and Wmc276 on chromosome 7B, and the map distance was 26.7 cM and 6.9 cM respectively. Sequence alignment, structure analysis, and comparative mapping all showed that TaCKX2 was highly homologous to OsCKX11 in rice. From the closely evolutionary relationship of orthologous CKX genes, we considered that CKX genes may be very conservative in cereals.

Key words: Wheat, Cytokinin oxidase/dehydrogenase, Gene cloning, Chromosomal localization, Genetic mapping