一个水稻“斑马叶”叶色突变体基因zebraleaf2(zl2)的图位克隆

doi:10.3969/j.issn.10017216.2013.03.002

中国水稻科学 ›› 2013, Vol. 27 ›› Issue (3): 231-239.DOI: 10.3969/j.issn.10017216.2013.03.002

一个水稻“斑马叶”叶色突变体基因zebraleaf2(zl2)的图位克隆

刘胜^{1, 2, #} ,魏祥进^{1, #} ,邵高能¹ ,唐绍清^{1, *} ,胡培松^{1, *}

¹中国水稻研究所国家水稻改良中心水稻生物学国家重点实验室, 浙江杭州 310006; ²杭州师范大学生命与环境科学学院, 浙江杭州 310036;

收稿日期:2012-11-20 修回日期:2013-01-25 出版日期:2013-05-10 发布日期:2013-05-10
通讯作者: 唐绍清1, * ,胡培松1, *
基金资助:
国家自然科学基金资助项目(31201193, 31201195); 浙江省自然科学基金资助项目(Y12C130012); 国家863计划资助项目(2011AA10A101); 中央级公益性科研院所基本科研业务费专项(2012RG0021)。

Mapbased Cloning of A ‘Zebra’ Leaf Mutant Gene zl2 in Rice

LIU Sheng ^{1, 2, #} , WEI Xiangjin ^{1, #} , SHAO Gaoneng¹, TANG Shaoqing ^{1, *} , HU Peisong ^{1, *}

¹State Key Laboratory of Rice Biology, Key Laboratory of Rice Biology and Breeding of Ministry of Agriculture, China National Rice Research Institute, Hangzhou 310006, China; ²College of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou 310036, China;

Received:2012-11-20 Revised:2013-01-25 Online:2013-05-10 Published:2013-05-10
Contact: TANG Shaoqing1, *, HU Peisong1, *

摘要/Abstract

摘要： 从粳稻品种Asominori组培后代中获得一个稳定遗传的黄绿相间叶色突变体（zebra leaf 2， zl2）。该突变体在苗期表现为黄绿相间的斑马状，分蘖后期斑马叶性状逐渐减弱，到抽穗期叶片逐渐变为淡黄色。与野生型相比，zl2 在3叶期、分蘖盛期、抽穗期及成熟期叶片的叶绿素、类胡萝卜素含量显著降低，成熟后其结实率、千粒重、株高也显著下降。电镜观察结果显示，苗期zl2叶片黄色部分叶肉细胞中叶绿体显微结构发生了明显的异常，而绿色部分与野生型基本一致。遗传分析结果表明，zl2突变性状受一对隐性核基因控制。从zl2与籼稻品种南京11衍生的F2群体中挑选1607株表现为突变性状的分离单株，最终将该突变基因定位于第11染色体约164.3 kb的区域内。基因预测表明该区域内存在13个ORFs，其中ORF12编码一个类胡萝卜素异构酶，序列分析表明突变体中的该基因第10个内含子与第11外显子的交界处碱基A突变为T，导致cDNA发生错误剪切，缺失4个碱基，产生移码突变，并于第395个氨基酸处提前终止。RTPCR分析表明，相对野生型在突变体中ZL2的表达量显著下降，同时叶色相关基因PORA、RbcL、RbcS、 Cab1、Cab2、 psaA、psbA、OsDVR表达量也显著下降，而HEMA1、YGL1、V1、V2、SPP、OsPPR的表达量显著上升。结果表明ZL2在水稻叶绿素合成及叶绿体发育中起着重要作用。

关键词: 叶色突变体;遗传分析, 图位克隆, 表达分析, 水稻

Abstract: A yellow and green zebra leaf mutant zebraleaf 2 (zl2) displayed a distinctive zebra leaf at seedling stage, and the leaves of the mutant gradually turned yellow after heading. Compared with wild type, the chlorophyll and carotenoid contents of zl2 were significantly reduced at trileaf stage, tillering stage, heading stage and mature stage. The seed setting rate, grain weight and plant height of zl2 also decreased significantly. The ultrastructures of chloroplasts in the mesophyll cells of the yellow part of seedling of zl2 have obvious differences from that of wild type. Genetic analysis showed that the phenotype of zl2 was controlled by a single recessive nuclear gene. Total 1607 zebra leaf plants selected from the F2 population of zl2 / Nanjing 11 were used to finemap the mutant gene. Finally, ZL2 gene was narrowed to the 164.3 kb region on chromosome 11. There are 13 ORFs in this region, and a single base mutation was found in the ORF12 which encode a carotenoid isomerase. And as a result, four bases were missed in cDNA of ORF12 of zl2, leading to frameshift and the amino acid premature termination at the 395th. Compared with the wild type, the expression level of ZL2 and the genes involved in the control of leaf color and photosynthesis, such as PORA, RbcL, RbcS， Cab1， Cab2， psaA, psbA and OsDVR decreased significantly in the mutant, while those of HEMA1, YGL1, V1, V2, SPP and OsPPR increase significantly. The results suggested that ZL2 act a very important role in chlorophyll biosynthesis, chloroplast development and photosynthesis in rice.

Key words: leaf mutant, genetic analysis, mapbased cloning, express analysis, rice

中图分类号:

刘胜1, 2, # ,魏祥进1, # ,邵高能1 ,唐绍清1, * ,胡培松1, *. 一个水稻“斑马叶”叶色突变体基因zebraleaf2(zl2)的图位克隆[J]. 中国水稻科学, 2013, 27(3): 231-239.

LIU Sheng1, 2, #, WEI Xiangjin1, #, SHAO Gaoneng1, TANG Shaoqing1, *, HU Peisong1, *. Mapbased Cloning of A ‘Zebra’ Leaf Mutant Gene zl2 in Rice[J]. Chinese Journal of Rice Science, 2013, 27(3): 231-239.

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一个水稻“斑马叶”叶色突变体基因zebraleaf2(zl2)的图位克隆

Mapbased Cloning of A ‘Zebra’ Leaf Mutant Gene zl2 in Rice

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