中国农业科学 ›› 2017, Vol. 50 ›› Issue (9): 1551-1558.doi: 10.3864/j.issn.0578-1752.2017.09.001

• 作物遗传育种·种质资源·分子遗传学 •    下一篇

水稻矮化剑叶卷曲突变体dcfl1的鉴定与基因精细定位

张孝波,谢佳,张晓琼,田维江,何沛龙,刘思岑,何光华,钟秉强,桑贤春   

  1. 西南大学水稻研究所/转基因植物与安全控制重庆市重点实验室,重庆 400715
  • 收稿日期:2016-11-09 出版日期:2017-05-01 发布日期:2017-05-01
  • 通讯作者: 桑贤春,E-mail:sangxianchun@163.com
  • 作者简介:张孝波,E-mail:zhangxiaobo@163.com。
  • 基金资助:
    中央高校基本科研业务费项目(XDJK2013A023)、重庆市研究生科研创新项目(CYS14043)

Identification and Gene Mapping of a Dwarf and Curled Flag Leaf Mutant dcfl1 in Rice (Oryza sativa L.)

ZHANG XiaoBo, XIE Jia, ZHANG XiaoQiong, TIAN WeiJiang, HE PeiLong, LIU SiCen, HE GuangHua, ZHONG BingQiang, SANG XianChun   

  1. Rice Research Institute of Southwest University/Chongqing Key Laboratory of Application and Safety Control of Genetically Modified Crops, Chongqing 400715
  • Received:2016-11-09 Online:2017-05-01 Published:2017-05-01

摘要:

【目的】对一个水稻矮化剑叶卷曲突变体进行鉴定与基因定位,为水稻等禾谷类作物剑叶形态发育及分子改良奠定基础。【方法】在籼型水稻恢复系缙恢10号的甲基磺酸乙酯(EMS)突变库中筛选到一个隐性矮化剑叶卷曲突变体,命名为dcfl1(dwarf and curled flag leaf 1)。田间小区种植,全生育期内观察dcfl1和野生型的株型变化。苗期利用扫描电镜观察叶鞘内表皮细胞大小;孕穗期和抽穗期利用石蜡切片观察剑叶基部形态;开花期测定剑叶、倒2叶和倒3叶的叶绿素含量;成熟期考查株高、有效穗数、穗实粒数、结实率和千粒重等主要农艺性状。配制西农1A/dcfl1杂交组合,利用F1和F2群体进行遗传分析,并利用F2隐性群体进行基因定位。【结果】生育期内,突变体dcfl1都表现出矮化性状。dcfl1叶鞘内表皮细胞长度明显比野生型要短,达到了极显著水平。与野生型相比,穗长、倒1节间和倒2节间均显著变短,倒3节间和倒4节间无显著变化。抽穗期dcfl1剑叶的叶片和叶鞘连接处硬化,剑叶基部展开受阻,半边叶片向内卷曲,剑叶上部和中部正常,其他叶片也正常。农艺性状调查发现,dcfl1的有效穗数为14.24,极显著高于野生型的11.62,穗粒数、实粒数、结实率和千粒重等则无显著变化。此外,dcfl1的叶色略深,剑叶、倒2叶和倒3叶的叶绿素a含量均极显著高于野生型,类胡萝卜素含量也略有升高,但仅剑叶达到极显著差异水平,叶绿素b的含量则无显著变化。西农1A/dcfl1的F1群体中,株高和剑叶表型与野生型一致。F2群体中分离出正常和突变两种表型,突变表型与dcfl1类似,植株株高变矮,剑叶基部特异卷曲,说明矮化和剑叶基部特异卷曲是一对共分离性状。且两种表型分离比符合3﹕1,表明dcfl1突变型受1对隐性核基因控制。利用620株F2隐性单株,最终将DCFL1精细定位在第3染色体短臂InDel标记Ind03-11和Ind03-6之间78 kb的物理范围内,包含15个注释基因,为DCFL1的克隆和水稻剑叶形态发育机理研究奠定了基础。【结论】dcfl1是一个水稻矮化剑叶基部特异卷曲突变体,基因精细定位在第3染色体78 kb的物理范围内。

关键词: 水稻, 矮化, 剑叶卷曲, 遗传分析, 精细定位

Abstract:

【Objective】Leaf blade is an important factor of plant type, which is directly related to leaf photosynthetic area and light energy utilization. Flag leaf is most prominently in the formation of rice production. Study of the genes which regulate flag leaf development in rice is of very significance in rice functional genomics research and molecular breeding. A novel flag leaf mutant has been identified and the results of study will provide a foundation for the research of leaf morphological formation and plant type breeding in Oryza sativa L.【Method】A dwarf and curled flag leaf mutant (dcfl1) was discovered from the progeny of indica restorer line Jinhui10 with seeds treated by ethyl methane sulfonate (EMS) and the traits of dwarf and curled flag leaf base inherited steadily after multi-generations’ self-fertility. The second leaf sheath was observed by scanning electron microscopy (SEM) at the three-leaf stage. The flag leaf base was used for paraffin section at the booting and heading stages. At the blooming stage, the characteristics of chloroplast pigment of the flag, second and third leaf blades were measured. At the maturity stage, agronomic traits such as plant height, panicle length, efficient panicle per plant, seed number per panicle, filled grain number per panicle, seed setting ratio, and 1000-seed weight were measured. The dcfl1 was crossed with indica sterile line Xinong 1A, and the F1 and F2 generations were used for genetic analysis. Additionally, gene mapping was performed based on the recessive individuals of the F2 generation of Xinong 1A/ dcfl1.【Result】The dcfl1 was dwarf in all phases of plant development. The cell length of the 2nd leaf sheath surface of the dcfl1 was significantly shorter than the wild type. The panicle length, the first and the second internode of the dcfl1 were all significantly shorter than those of the wild type. The dcfl1 displayed a severe curl at the base of flag leaf blade after the heading stage, while the upper of flag leaf blade was nearly normal in the flag leaf. Meanwhile, the other leaf blades appeared as normal as the wild type. No significant differences were detected in grain number per panicle, filled grain number per panicle, seed setting rate and 1000-seed weight between the dcfl1 and the wild type. However, the number of the tiller in the dcfl1 was more than the wild type and the efficient panicle per plant was increased significantly than the wild type. Having the dark green leaves, the contents of chlorophyll a and total chlorophyll in the dcfl1 increased significantly compared with those of the wild type for the flag leaves, the second and the third leaves. Genetic analysis indicated that the dwarf and curled flag leaf traits of dcfl1 were controlled by a recessive nuclear gene. Based on the F2 population derived from a cross between the dcfl1 and an indica sterile line, Xinong 1A, the gene was fine mapped on chromosome 3 between InDel marker Ind03-11 and Ind03-6 with the physical distance 78 kb, containing fifteen annotated genes.【Conclusion】The dcfl1 is a novel recessive dwarf and curled flag leaf mutant coming from EMS-inducement. The DCFL1 was mapped on chromosome 3 with 78 kb physical distance. These results will provide a foundation for map-based cloning of DCFL1 gene and understanding of the molecular mechanism of the rice flag leaf.

Key words: rice (Oryza sativ L.), dwarf, curled flag leaf, genetic analysis, gene mapping