拟南芥突变体cka3-2的筛选及其功能初步研究
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摘要
CKA3-2基因属于拟南芥CK基因家族成员,该基因编码一条691个氨基酸残基的多肽.利用三引物法和实时定量PCR法鉴定获得CKA3-2基因对应的T-DNA插入纯合突变体cka3-2,并对其表T型变化进行了观察.实时定量PCR分析表明:CKA3-2基因在茎生叶中表达最高,花和茎中次之,在根、莲座叶和果荚中表达量较低.赤霉素使CKA3-2基因的表达升高,6 h时表达量明显增加,8 h时表达量下降.用50μmol/L GA3处理Col-0和cka3-2幼苗6 h,结果发现赤霉素信号转导通路相关基因如KS、KAO1、KAO2、GA2ox1、GA3OX1和GA20OX1等在两种幼苗中的相对表达量发生明显变化.此外,突变体cka3-2相对野生型Col-0来说,具有较明显的早花现象.
CKA3-2 gene is a member of CK(Casein kinase) gene family, and this gene codes 691 anmino acids. Three primer PCR method and real-time PCR technique were used to identify the homozygous T-DNA mutant cka3-2 of CKA3-2 gene in Arabidopsis. The phenotypic changes of cka3-2 and Col-0 plants were observed. The results of real-time PCR analysis showed that the CKA3-2 gene was highly expressed in stem leaf, flower and stem, and less in root, rosette leaf and silique. The expression of CKA3-2 gene was induced by GA3, and its expression was significantly increased at 6 h. However, its expression was decreased at 8 h.The Col-0 and cka3-2 mutant seedlings were treated with 50 μmol/L GA3for 6 h, and obvious changes were found for the expression levels of related genes involved in GA signal transduction pathways including KS,KAO1, KAO2, GA2ox1, GA3OX1 and GA20OX12. Furthermore, cka3-2 mutants flowered early compared to Col-0 plants.
CKA3-2 gene is a member of CK(Casein kinase) gene family, and this gene codes 691 anmino acids. Three primer PCR method and real-time PCR technique were used to identify the homozygous T-DNA mutant cka3-2 of CKA3-2 gene in Arabidopsis. The phenotypic changes of cka3-2 and Col-0 plants were observed. The results of real-time PCR analysis showed that the CKA3-2 gene was highly expressed in stem leaf, flower and stem, and less in root, rosette leaf and silique. The expression of CKA3-2 gene was induced by GA3, and its expression was significantly increased at 6 h. However, its expression was decreased at 8 h.The Col-0 and cka3-2 mutant seedlings were treated with 50 μmol/L GA3for 6 h, and obvious changes were found for the expression levels of related genes involved in GA signal transduction pathways including KS,KAO1, KAO2, GA2ox1, GA3OX1 and GA20OX12. Furthermore, cka3-2 mutants flowered early compared to Col-0 plants.
引文
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[2]JOHNSON L N,LOWE E D,NOBLE M E M,et al.The structural basis for substrate recognition and control by protein kinases[J].FEBS Letters,1998,1(430):1-11.
[3]GIETZEN K F,VIRSHUP D M.Identification of inhibitory autophosphorylation sites in casein kinase I epsilon[J].Journal of Biological Chemistry,1999,274(45):32063-32070.
[4]AKASHI M,TSUCHIYA Y,YOSHINO T,et al.Control of intracellular dynamics of mammalian period proteins by casein kinase I epsilon(CKIepsilon)and CKIdelta in cultured cells[J].Molecular and Cellular Biology,2002,22(6):1693-1703.
[5]EIDE E J,VIELHABER E L,HINZ W A,et al.The circadian regulatory proteins BMAL1 and cryptochromes are substrates of casein kinase Iepsilon[J].Journal of Biology Chemistry,2002,277(19):17248-17254.
[6]MCKAY R M,PETERS J M,GRAFF J M.The casein kinase I family in Wnt signaling[J].Development Biology,2001,235(2):388-396.
[7]喻达时,赵琼,邓克勤,等.拟南芥CK1A基因功能初步研究[J].遗传(YU Da-shi,ZHAO Qiong,DENG Ke-qin,et al.Preliminary studies on the function of Arabidopsis CK1A gene[J].Hereditas,2009,31(10):1037-1042.
[8]郭新红,王婕,秦玉芝,等.拟南芥CKL3基因表达对非生物胁迫的响应[J].植物生理学通讯(GUO Xin-hong,WANG Jie,QING Yu-zhi,et al.Responses of CKL3 gene expression to abiotic stresses in Arabidopsis[J].Plant Physiology Communications,2008,44(4):673-676.
[9]LIU W,XU Z H,LUO D,et al.Roles of OsCKI1,a rice casein kinase I,in root development and plant hormone sensitivity[J].Plant Journal,2003,36(2):189-202.
[10]PINNA L A,MEGGIO F.Protein kinase CK2“casein kinase-2”and its implication in cell division and proliferation[J].Progress in Cell Cycle Research,1997,24(3):77-97.
[11]DANIEL X,SHOJ I,SUGANO S,et al.CK2 phosphorylation of CCA1 is necessary for circadian oscillator function in Arabidopsis[J].Proceedings of the National Academy of Sciences,2004,101(16):3292-3297.
[12]PERALES M,PORTOL魪S S,M魣S P.The proteasome-dependent degradation of CKB4 is regulated by the Arabidopsis biological clock[J].Plant Journal,2006,46(5):849-860.
[13]汪启明.拟南芥DBB亚家族基因表达的光调控和DBB1α基因功能分析[D].长沙:湖南大学(WANG Qi-ming.Light Regulated Expression of DBB Subfamily Genes and Functional Study on DBB1a in Arabidopsis[D].Changsha:Hunan University,2009.1-23.