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光敏色素B基因(PHYB)启动子突变及与大白菜开花时间的关联分析
刘栓桃,张志刚,李巧云,王淑芬,赵智中,卢金东,张晓燕,徐文玲,刘贤娴,付卫民
山东省农业科学院蔬菜花卉研究所
Association Analysis Between Phytochrome B Gene(PHYB) Promoter Mutantions and Flowering Time in Chinese cabbage(Brassica rapa L. ssp. pekinensis)
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摘要 光敏色素B(phytochrome B, PHYB)是植物光周期依赖开花途径的关键调控因子之一。本研究对极端晚花和早花大白菜(Brassica rapa L. ssp. pekinensis)自交系06-247与He102的PHYB全基因组序列进行了生物信息学分析。结果表明,二者全长分别是5 986和5 528 bp,均与数据库中Chiifu-401的同源序列有不同程度的差异,表明二者均是大白菜PHYB基因的突变体,并命名为BraphyB1(GenBank登录号: KJ866947)和BraphyB2(GenBank登录号: KJ866948)。二者之间有476个插入/缺失(Inserts/Deletions, InDels)和57个SNPs。启动子区有473个InDels和31个SNPs、编码区有3个InDels和26个SNPs。InDels主要体现为phyB2在启动子区、5'非翻译区和编码区分别缺失445、18和3 bp。phyB2启动子缺失的445 bp中包含两个TATABOX4和两处成髓细胞血症 (myeloblastosis, MYB)类转录因子结合基序,一处GA响应元件。开发了鉴定phyB1/phyB2中445 bp InDel突变的共显性标记 phyB1-762/phyB2-317,用该标记检测了以06-247与He102为亲本杂交构建的F2群体,利用F2代群体植株对抽薹开花时间与标记进行了关联分析,结果表明,phyB1-762/phyB2-317分别与晚花/早花表型显著关联(P<0.05)。荧光定量RT-PCR显示,PHYB基因在He102各个发育阶段的表达水平均显著低于06-247(P<0.01)。上述结果表明,大白菜PHYB基因启动子InDel是造成PHYB基因表达差异和开花期改变的主要原因之一。本研究为系统研究PHYB基因在转录、翻译、蛋白结构等水平突变对大白菜开花时间影响的分子机制提供了线索。
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刘栓桃
张志刚
李巧云
王淑芬
赵智中
卢金东
张晓燕
徐文玲
刘贤娴
付卫民
关键词 大白菜光周期依赖开花期PHYB启动子突变    
Abstract:Phytochrome B (PHYB) is one of key factors regulating photoperiod-dependent flowering in plant. At present study, whole genomic sequences of PHYB, which were isolated from extremely late- and early-bolting Chinese cabbage (Brassica rapa L. ssp. pekinensis) inbred lines 06-247 and He102, were analyzed bioinformatically. The results revealed that the total length of PHYB in both lines were 5 986 and 5 528 bp respectively. And the sequences of both were not completely identical to that of Chiifu-401 from BRAD (Brassica Database), thus they were all mutants of wild type PHYB and named as BraphyB1(GenBank No. KJ866947) (from 06-247) and BraphyB2 (GenBank No. KJ866948)(from He102). There were 476 Inserts/Deletions(InDels) and 57 SNPs, among which 473 InDels and 31 SNPs located in promotor and 3 InDels and 26 SNPs located in coding region. InDels appeared 445, 18 and 3 bp deletions in phyB2 promoter, 5'UTR and coding region, respectively, among which 445 bp deletion in promotor containing two TATABOX4, two MYB-like transcription factors biding sites and one GA-responsive element; 18 bp deletion in 5'UTR might influence secondary structure of mature mRNA and then the scanning of pre-initiation complex and translation efficiency; 3 bp deletion in coding region of phyB2 caused an amino acid deletion. Furtermore, among 26 SNPs in coding region, 3 were nonsynonymous mutations which caused amino acid subtitutions between phyB1 and phyB2. A set of codorminant markers relating to large fragment InDel in promotor were developed and named as phyB1-762/phyB2-317. A segregating F2 population derived from 06-247 and He102 was detected with the markers. Association analysis between markers and flowering time revealed that phyB1-762 and phyB2-317 were significantly (P<0.05) related to late and early flowering respectively. Real-time quantitative RT-PCR analysis revealed that the relative mRNA level in tissues of He102 were evidantly lower than that in 06-247, the difference was statistically the most significant (P<0.01) at all development stages tested. Thus the large fragment deletion mutation in promotor was one of the most important factors causing low level phyB2 expresson and early flowering in He102. The results provide clues for research on molecular mechanism which influences Chinese cabbage flowering time by PHYB mutation at the level of trancription, translation and protein structure.
Key wordsBrassica rapa L. ssp. pekinensis    Photoperiod-dependent    Flowering time    PHYB promoter    Mutation
收稿日期: 2013-12-12     
ZTFLH:  S634.1  
通讯作者: 赵智中   
引用本文:   
刘栓桃,张志刚,李巧云,王淑芬,赵智中,卢金东,张晓燕,徐文玲,刘贤娴,付卫民. 光敏色素B基因(PHYB)启动子突变及与大白菜开花时间的关联分析[J]. , 2014, 22(7): 853-861.
链接本文:  
http://journal05.magtech.org.cn/Jwk_ny/CN/     或     http://journal05.magtech.org.cn/Jwk_ny/CN/Y2014/V22/I7/853
 
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