甘蓝型油菜BnEOD3基因克隆与比较测序分析
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摘要
【目的】克隆拟南芥籽粒大小发育母体效应基因EOD3在甘蓝型油菜籽粒中表达的同源基因拷贝,并在大、小籽粒材料间开展序列比较分析。【方法】以1份大籽粒与1份小籽粒种质发育中的籽粒为研究材料,利用RT-PCR克隆发育中籽粒表达的BnEOD3基因拷贝,结合生物信息学手段在大、小籽粒材料间进行差异比较分析。【结果】生物信息学分析发现BnEOD3基因在甘蓝型油菜中有4个同源拷贝(BnaC04g00760D,BnaA05g01200D,BnaC04g50960D,BnaA04g27100D)。其中BnaA04g27100D在大、小籽粒材料发育籽粒中均有表达,且在第114碱基处存在SNP变异(大籽粒为C,小籽粒为G),BnaC04g50960D则仅在小籽粒材料中检测到表达,BnaC04g00760D与BnaA05g01200D基因则在大、小籽粒中均未检测到表达。【结论】BnEOD3基因仅有2个拷贝(BnaA04g27100D和BnaC04g50960D)在发育籽粒中检测到表达,其中BnaA04g27100D在第114碱基处的SNP变异导致氨基酸序列甘氨酸(G)与丙氨酸(A)的氨基酸的差异,可能与大、小籽粒性状有关。
【Objective】BnEOD3,the homologous gene of EOD3 controlling the seed development in arabidopsis,were cloned and comparative analyzed. 【Method】The developing seeds obtained from 1 lager rape seed germplasm and 1 normal rape seed germplasm were used as research materials to clone BnEOD3 with RT-PCR technique. The expressed copy of BnEOD3 and the copy sequences were investigated and used for comparative analysis between the extra lager rape seed germplasm and small rape seed germplasm.【Result】There were 4 copies of BnEOD3,BnaC04 g00760 D,BnaA05 g01200 D,BnaC04 g50960 D and BnaA04 g27100 D,existing in rape genomes,and only 2 copy,BnaA04 g27100 D and BnaC04 g50960 D,were detected expressing in developing seed. Furthermore,the differences of the number of expressed copy and 1 SNP site were also observed between the larger and normal seed materials.【Conclusion】There was one SNP difference at the 114 bp of BnaA04 g27100 D between the larger and normal seed materials,which might be related with difference of seed size.
【Objective】BnEOD3,the homologous gene of EOD3 controlling the seed development in arabidopsis,were cloned and comparative analyzed. 【Method】The developing seeds obtained from 1 lager rape seed germplasm and 1 normal rape seed germplasm were used as research materials to clone BnEOD3 with RT-PCR technique. The expressed copy of BnEOD3 and the copy sequences were investigated and used for comparative analysis between the extra lager rape seed germplasm and small rape seed germplasm.【Result】There were 4 copies of BnEOD3,BnaC04 g00760 D,BnaA05 g01200 D,BnaC04 g50960 D and BnaA04 g27100 D,existing in rape genomes,and only 2 copy,BnaA04 g27100 D and BnaC04 g50960 D,were detected expressing in developing seed. Furthermore,the differences of the number of expressed copy and 1 SNP site were also observed between the larger and normal seed materials.【Conclusion】There was one SNP difference at the 114 bp of BnaA04 g27100 D between the larger and normal seed materials,which might be related with difference of seed size.
引文
[1]王璐.中国油菜产业安全研究[D].武汉:华中农业大学博士学位论文,2014:24-27.
[2]Fang W,Wang Z,Cui R,et al. Maternal control of seed size byEOD3/CYP78A6 in Arabidopsis thaliana[J]. Plant J,2012,70(6):929-939.
[3]余世聪,梁浩,李壮.高油酸含量甘蓝型油菜BnaLCR78基因的克隆、表达及过表达载体的构建[J].西南农业学报,2018,31(1):6-13.
[4]祝利霞,张冬晓,傅廷栋,等. 20年来中国冬油菜新品种产量和抗病性状分析[J].中国农学通报,2010,24:375-380.
[5]Wang X,Chen L,Wang A,et al. Quantitative trait loci analysis andgenome-wide comparison for silique related traits in Brassica napus[J]. Bmc Plant Biol,2016,16:71.
[6]Liu J,Hua W,Hu Z,et al. Natural variation in ARF18 gene simul-taneously affects seed weight and silique length in polyploid rapeseed[J]. Proceedings of the National Academy of Sciences of the UnitedStates of America,2015,112(37):5123-5132.
[7]Yang P,Shu C,Chen L,et al. Identification of a major QTL for si-lique length and seed weight in oilseed rape(Brassica napus L.)[J]. Theoretical and Applied Genetics,2012,125(2):285-296.
[8]Fan C,Cai G,Qin J,et al. Mapping of quantitative trait loci anddevelopment of allele-specific markers for seed weight in Brassica na-pus[J]. Theoretical and Applied Genetics,2010,121(7):1289-1301.
[9]Shi J,Li R,Qiu D,et al. Unraveling the complex trait of crop yieldwith quantitative trait loci mapping in Brassica napus[J]. Genetics,2009,182(3):851-861.
[10]Sundaresan V. Control of seed size in plants[J]. Proceedings of theNational Academy of Sciences of the United States of America,2005,102(50):17887-17888.
[11]Dante R A,Larkins B A,Sabelli P A. Cell cycle control and seeddevelopment[J]. Front Plant Sci,2014(5):493.
[12]Li N,Li Y. Ubiquitin-mediated control of seed size in plants[J].Front Plant Sci.,2014(5):332.
[13]Radchuk V,Borisjuk L. Physical,metabolic and developmentalfunctions of the seed coat[J]. Front Plant Sci,2014(5):510.
[14]Meinke D W,Cherry J M,Dean C,et al. Arabidopsis thaliana:amodel plant for genome analysis[J]. Science,1998,282(5389):679-682.
[15]Cai G,Yang Q,Yi B,et al. A complex recombination pattern inthe genome of allotetraploid Brassica napus as revealed by a high-den-sity genetic map[J]. Plos One,2014,9(10):e109910.
[16]李静.甘蓝型油菜千粒重及相关性状QTL定位[D].武汉:华中农业大学硕士学位论文,2011:14-15.
[17]陈苇,李劲峰,和爱花,等.甘蓝型油菜特大籽粒新种质创制及评价[J].中国油料作物学报,2011(5):513-516.
[18]陈苇,李劲峰,张国建,等.特大粒甘蓝型油菜籽粒和角果发育形态特征[J].中国油料作物学报,2013(6):658-664.
[19]Du L,Li N,Chen L,et al. The ubiquitin receptor DA1 regulatesseed and organ size by modulating the stability of the ubiquitin-spe-cific protease UBP15/SOD2 in Arabidopsis[J]. Plant Cell,2014,26(2):665-677.
[20]Li Y,Zheng L,Corke F,et al. Control of final seed and organ sizeby the DA1 gene family in Arabidopsis thaliana[J]. Genes&Devel-opment,2008,22(10):1331-1336.
[21]Zhu Q,Li B,Mu S,et al. TTG2-regulated development is relatedto expression of putative AUXIN RESPONSE FACTOR genes in to-bacco[J]. BMC Genomics,2013,14:806.
[22]Ohto M A,Fischer R L,Goldberg R B,et al. Control of seed massby APETALA2[J]. Proceedings of the National Academy of Sciencesof the United States of America,2005,102(8):3123-3128.
[2]Fang W,Wang Z,Cui R,et al. Maternal control of seed size byEOD3/CYP78A6 in Arabidopsis thaliana[J]. Plant J,2012,70(6):929-939.
[3]余世聪,梁浩,李壮.高油酸含量甘蓝型油菜BnaLCR78基因的克隆、表达及过表达载体的构建[J].西南农业学报,2018,31(1):6-13.
[4]祝利霞,张冬晓,傅廷栋,等. 20年来中国冬油菜新品种产量和抗病性状分析[J].中国农学通报,2010,24:375-380.
[5]Wang X,Chen L,Wang A,et al. Quantitative trait loci analysis andgenome-wide comparison for silique related traits in Brassica napus[J]. Bmc Plant Biol,2016,16:71.
[6]Liu J,Hua W,Hu Z,et al. Natural variation in ARF18 gene simul-taneously affects seed weight and silique length in polyploid rapeseed[J]. Proceedings of the National Academy of Sciences of the UnitedStates of America,2015,112(37):5123-5132.
[7]Yang P,Shu C,Chen L,et al. Identification of a major QTL for si-lique length and seed weight in oilseed rape(Brassica napus L.)[J]. Theoretical and Applied Genetics,2012,125(2):285-296.
[8]Fan C,Cai G,Qin J,et al. Mapping of quantitative trait loci anddevelopment of allele-specific markers for seed weight in Brassica na-pus[J]. Theoretical and Applied Genetics,2010,121(7):1289-1301.
[9]Shi J,Li R,Qiu D,et al. Unraveling the complex trait of crop yieldwith quantitative trait loci mapping in Brassica napus[J]. Genetics,2009,182(3):851-861.
[10]Sundaresan V. Control of seed size in plants[J]. Proceedings of theNational Academy of Sciences of the United States of America,2005,102(50):17887-17888.
[11]Dante R A,Larkins B A,Sabelli P A. Cell cycle control and seeddevelopment[J]. Front Plant Sci,2014(5):493.
[12]Li N,Li Y. Ubiquitin-mediated control of seed size in plants[J].Front Plant Sci.,2014(5):332.
[13]Radchuk V,Borisjuk L. Physical,metabolic and developmentalfunctions of the seed coat[J]. Front Plant Sci,2014(5):510.
[14]Meinke D W,Cherry J M,Dean C,et al. Arabidopsis thaliana:amodel plant for genome analysis[J]. Science,1998,282(5389):679-682.
[15]Cai G,Yang Q,Yi B,et al. A complex recombination pattern inthe genome of allotetraploid Brassica napus as revealed by a high-den-sity genetic map[J]. Plos One,2014,9(10):e109910.
[16]李静.甘蓝型油菜千粒重及相关性状QTL定位[D].武汉:华中农业大学硕士学位论文,2011:14-15.
[17]陈苇,李劲峰,和爱花,等.甘蓝型油菜特大籽粒新种质创制及评价[J].中国油料作物学报,2011(5):513-516.
[18]陈苇,李劲峰,张国建,等.特大粒甘蓝型油菜籽粒和角果发育形态特征[J].中国油料作物学报,2013(6):658-664.
[19]Du L,Li N,Chen L,et al. The ubiquitin receptor DA1 regulatesseed and organ size by modulating the stability of the ubiquitin-spe-cific protease UBP15/SOD2 in Arabidopsis[J]. Plant Cell,2014,26(2):665-677.
[20]Li Y,Zheng L,Corke F,et al. Control of final seed and organ sizeby the DA1 gene family in Arabidopsis thaliana[J]. Genes&Devel-opment,2008,22(10):1331-1336.
[21]Zhu Q,Li B,Mu S,et al. TTG2-regulated development is relatedto expression of putative AUXIN RESPONSE FACTOR genes in to-bacco[J]. BMC Genomics,2013,14:806.
[22]Ohto M A,Fischer R L,Goldberg R B,et al. Control of seed massby APETALA2[J]. Proceedings of the National Academy of Sciencesof the United States of America,2005,102(8):3123-3128.