Shen Y G, Du B X, Zhang J S, Chen S Y
Laboratory of Plant Biotechnology, Institute of Genetics, Chinese Academy of Sciences, Beijing 100101, China.
Sheng Wu Gong Cheng Xue Bao. 2001 Jan;17(1):1-6.
Glycine betaine is a widespread osmopretectant existed in many organisms. In higher plant, glycine betaine is synthesized via a two-sep oxidation reaction: choline-->betaine aldehyde-->glycine betaine. The first step, also the speed-limiting step, is catalyzed by choline monooxygenase(CMO). Choosing halophyte Atriplex hortensis as material, we constructed a salt stress-induced cDNA library, and isolated a 1.77 kb length cDNA clone with spinach CMO cDNA as probe. The sequencing result showed a complete Open Reading Frame encoding a 438-amino-acid polypeptide which was 81% and 72% identified to CMO sequences of spinach and sugar beet in amino acid homology respectively. Compared with the CMO from spinach and sugar beet, the AhCMO had one conserved Rieske-Type [2Fe-2S] cluster-binding region and one conserved mononuclear Fe-binding motif. The expression pattern of AhCMO under salt stress was also stuied, the transcriptional level of AhCMO raised about three folds after the plant was treated with brine for 4 days. The AhCMO was then transfered into tobacco(Nictiana tabacum var. Xanthi) with 35S promotor and seven transgenic plants were certified by northern blot, these plants displayed some salt- and drought-stress tolerance when grew well on MS medium contained 1.2% NaCl or 10% PEG while the control was stagnated under the same cndition.
甘氨酸甜菜碱是一种广泛存在于许多生物体中的渗透保护剂。在高等植物中,甘氨酸甜菜碱通过两步氧化反应合成:胆碱→甜菜碱醛→甘氨酸甜菜碱。第一步也是限速步骤,由胆碱单加氧酶(CMO)催化。以盐生植物盐地碱蓬为材料,构建了盐胁迫诱导的cDNA文库,并以菠菜CMO cDNA为探针分离得到一个1.77 kb长的cDNA克隆。测序结果显示有一个完整的开放阅读框,编码一个438个氨基酸的多肽,其氨基酸同源性分别与菠菜和甜菜的CMO序列有81%和72%的一致性。与菠菜和甜菜的CMO相比,盐地碱蓬CMO有一个保守的Rieske型[2Fe-2S]簇结合区域和一个保守的单核铁结合基序。还研究了盐地碱蓬CMO在盐胁迫下的表达模式,用盐水处理植物4天后,盐地碱蓬CMO的转录水平提高了约三倍。然后将盐地碱蓬CMO与35S启动子一起转入烟草(烟草品种Xanthi),通过Northern杂交鉴定出7株转基因植株,这些植株在含有1.2% NaCl或10% PEG的MS培养基上生长良好时表现出一定的耐盐和耐旱胁迫能力,而对照在相同条件下生长停滞。