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作物学报 ›› 2010, Vol. 36 ›› Issue (07): 1067-1074.doi: 10.3724/SP.J.1006.2010.01067

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

甘蓝KAPP编码基因的克隆与序列分析

吴韦铷1,朱利泉1,*,李成琼3,杨昆1,唐章林2,任雪松3,王小佳3,*   

  1. 1西南大学植物生理生化实验室；2重庆市油菜工程中心；3西南大学蔬菜学实验室，重庆400716
  • 收稿日期:2010-02-01 修回日期:2010-04-19 出版日期:2010-07-12 网络出版日期:2010-05-20
  • 通讯作者: 朱利泉,E-mail: zhuliquan@swu.edu.cn; wxj@swu.edu.cn
  • 基金资助:

    本研究由国家自然科学基金项目（30671429，30971849）资助。

Cloning and Sequence Analysis of KAPP Gene in Brassica oleracea

WU Wui-Ru1,ZHU Li-Quan1*,LI Cheng-Qiong3,YANG Kun1,TANG Zhang-Lin1,REN Xue-Song3,WANG Xiao-Jia3*   

  1. 1 Plant Physiology and Biochemistry Laboratory of Southwest University, Chongqing 400716, China; 2 Chongqing Rapeseed Technology Research Center, Chongqing 400716, China; 3 Key Laboratory in Olericulture of Chongqing, Southwest University, Chongqing 400716, China
  • Received:2010-02-01 Revised:2010-04-19 Published:2010-07-12 Published online:2010-05-20
  • Contact: ZHU Li-Quan,E-mail: zhuliquan@swu.edu.cn; wxj@swu.edu.cn

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摘要： 采用PCR、RT-PCR及其他分子生物学方法，以甘蓝基因组DNA、花蕾RNA和叶片RNA为模板，分别对甘蓝KAPP gDNA和KAPP cDNA进行扩增，分别获得3 247 bp的KAPP gDNA片段、1 699 bp的KAPP cDNA片段﹑1 578 bp的花蕾KAPP2 cDNA片段和1 581 bp的叶片KAPP2 cDNA片段。对克隆的甘蓝KAPP gDNA和cDNA(结合报道的KAPP cDNA)进行比对表明，甘蓝KAPP基因包含11个内含子，均符合“GU-AG”剪接规则，并且克隆得到的KAPP cDNA序列与报道的KAPP cDNA序列有6处单个碱基的差异，但两者的氨基酸序列完全一致。然而花蕾KAPP2 cDNA、叶片KAPP2 cDNA片段与报道的KAPP cDNA序列相似性分别为85.2%和85.0%。这两个序列分别在590 bp和593 bp处较早出现一个无义突变引起的终止密码子。Blast分析表明, 两基因编码的氨基酸序列与拟南芥KAPP氨基酸序列相似性最高，其次为甘蓝KAPP氨基酸序列。以已报道的8种植物KAPP的CDS及本实验所克隆的两个KAPP2序列构建分子进化树，获得序列与甘蓝KAPP序列聚为一支。结合比较作图及分子进化树，推测KAPP基因在甘蓝基因组上有两个拷贝，而笔者克隆到的KAPP2 cDNA序列是其中一个拷贝，是KAPP进化过程中突变失活的拟基因。

关键词: 自交不亲和性, 信号转导, KAPP基因, 甘蓝

Abstract: The gDNA and cdna fragments of KAPP gene were amplified from genomic DNA, bud Rna and leaf Rna in Brassica oleracea by PCR and RT-PCR and other molecular biology methods. We initially obtained a fragment of KAPP gDNA with length of 3 247 bp, a fragment of KAPP cDNA with length of 1 699 bp, as well as a bud cdna with length of 1 578 bp and a leaf cdna with length of 1 581 bp which were called KAPP2 cDNA of bud and KAPP2 cDNA of leaf respectively. Compared gDNA with cDNA of KAPP, we found that there were 11 introns in KAPP gene, and these introns all followed typical GU-AG rule. Between KAPP cDNA which we cloned and KAPP cDNA released there are six nucleotide differences, but they encode a same amino acid sequence. Sequences analysis of the KAPP2 cDNA which we cloned from bud cdna and leaf cdna in Brassica oleracea showed that they share 85.2% and 85.0% identity with the reported KAPP cDNA respectively.We also found that the nonsense mutation in 590 bp of KAPP2 of bud cDNA and 593 bp of KAPP2 of leaf cDNA led to the early appearance of termination codon, who’s Blast indicated that both of them shared more identity in Arabidopsis thaliana than in Brassica oleracea. Based on KAPP cDNA sequence of eight species released by NCBI and two sequences of KAPP2 cDNA cloned in this study, we constructed a phylogenetic tree of KAPP gene, which showed that the two KAPP2 sequences were in the same group with released KAPP cDNA in Brassica oleracea. Based on all the above analysis, as well as some view of the comparative mapping research, we speculated that KAPP gene may have more than one copy in the genome of Brassica oleracea,the KAPP2 sequence cloned in this study may be the other copies of KAPP gene. Moreover, they are likely to be the pseudogene which was inactivated by mutation during evolution process. It has an important significance for the deep research between KAPP and Self-incompatibility. The results will provide some new insights into the research of molecular mechanism in SI and molecular evolution in Brassica oleracea.

Key words: Self-incompatibility, Signal transduction, KAPP gene, Brassica oleracea

[1] Nou I S, Watanabe M, Isogai A, Hinata K. Comparison of S-alleles n S-glycoproteins between two wild populations of Brassica campestris in Turkey and Japan. Sex Plant Reprod, 1993, 6: 79–86
[2] Schopfer C R, Nasrallah M E, Nasrallah J B. The male determinant of self-incompatibility in Brassica. Science, 1999, 286: 1697–1700
[3] Stein J C, Howlett B, Boyes D C, Nasrallah M E, Nasrallah J B. Molecular cloning of a putative receptor protein kinase gene encoded at the self-incompatibility locus of Brassica oleracea. Proc Nat Acad Sci USA, 1991, 88: 8816–8820
[4] Cohen P T W. Novel protein serine/threonine phosphatases: variety is the spice of life. Trends Biochem Sci, 1997, 22: 245–251
[5] Stone J M, Collinge M A, Smith R D, Horn M A, Walker J C. Interaction of a protein phosphatase with an Arabidopsis serine-threonine receptor kinase. Science, 1994, 266: 793–795
[6] Torii K U. Receptor kinase activation and signal transduction in plants: An emerging picture. Curr Opin Plant Biol, 2000, 3: 361–367
[7] Clark S E, Williams R W, Meyerowitz E M. The CLAVATA1 gene encodes a putative receptor kinase that controls shoot and floral meristem size in Arabidopsis. Cell, 1997, 89: 575–585
[8] Braun D M, Stone J M, Walker J C. Interaction of the maize and Arabidopsis kinase interaction doamins with a subset of receptor-like protein kinases: implication for transmembrane signaling in plants. Plant J, 1997, 12: 83–95
[9] Park A R, Cho S K, Yun U J, Jin M Y, Lee S H, Sachetto-Martins G, Park O K. Interaction of the Arabidopsis receptor protein kinase Wak1 with a glycine-rich protein, AtGRP-3. Biol Chem, 2001, 276: 26688–26693
[10] Gómez-Gómez L, Bauer Z, Boller T. Both the extracellular leucine-rich repeat domain and the kinase activity of FLS2are required for flagellin binding and signaling in Arabidopsis. Plant Cell, 2001, 13: 1155–1163
[11] Li J, Wen J Q, Lease K A, Doke J T, Tax F E, Walker J C. BAK1, an Arabidopsis LRR receptor-like protein kinase, interacts with BRI1 and modulates brassinosteroid signaling. Cell, 2002, 110: 213–222
[12] Shah K, Russinova E, Gadella T W Jr, Willemse J, De Vries S C. The Arabidopsis kinase-associated protein phosphatase controls internalization of the somatic embryogenesis receptor kinase 1. Genes, 2002, 16: 1707–2003
[13] Cabrillac D, Cock J M, Dumas C, Gaude T. The S-locus receptorkinase is inhibited by thioredoxins and activated by pollen coat proteins. Nature, 2001, 410: 220–223
[14] Williams R W, Wilson J M, Meyerowitz E M. A possible role forkinase-associated protein phosphatase in the Arabidopsis CLAVATA1 signaling pathway. Proc Natl Acad Sci USA, 1997, 94: 10467–10472
[15] Stone J M, Trotochaud A E, Walker J C, Clark S E. Control of meristem development by CLAVATA1 receptor kinase and kinase-associated protein phosphatase interactions. Plant Physiol, 1998, 117: 1217–1225
[16] Li Y-Y(李媛媛), Fu Y-D(傅廷栋), Ma C-Z(马朝芝). Advances of comparative genomics in Brassica. Chin Bull Bot (植物学通报), 2007, 24(2): 200–207 (in Chinese with English abstract)
[17] Vanoosthuyse V, Tichtinsky G, Dumas C, Gaude T, Cock J M. Interaction of calmodulin, a sorting nexin and kinase-associated protein phosphatase with the Brassica oleracea S locus receptor kinase. Plant Physiol, 2003, 133: 919–929
[18] Zhao Y-B(赵永斌), Zhu L-Q(朱利泉), Wang X-J(王小佳). Cloning and sequences analysis of m locus protein kinase gene from Brassica oleracea. Acta Agric Sin (作物学报), 2006, 32(1): 46–50 (in Chinese with English abstract)
[19] Liu D(刘东), Zhu L-Q(朱利泉), Wang X-J(王小佳). Cloning and characterization of SRK-binding protein THL1 gene from Brassica oleracea L. in self-incompatibility signaling process. Acta Hort Sin (园艺学报), 2003, 30(1): 56−58 (in Chinese with English abstract)
[20] Liu D(刘东), Zhu L-Q(朱利泉) ,Wang X-J(王小佳). Cloning and sequences analysis of ARC1 protein gene, a substrate of SRKin SI signal transduction from Brassica oleracea. Acta Agron Sin (作物学报), 2004, 30(5): 427–431 (in Chinese with English abstract)
[21] Li J, Smith G P, Walker J C. Kinase interaction domain of kinase-associated protein phosphatase, a phosphoprotein binding domain. Proc Natl Acad Sci USA, 1999, 96: 7821−7826
[22] Braun D M, Walker J C. Plant transmembrane receptors: New pieces in the signaling puzzle. Trends Biochem Sci, 1996, 21: 70−73
[23] Philip W B. Receptor kinases in plant development. Trends Plant Sci, 1998, 3: 384−388
[24] Nowak M A, Boerlijst M C, Cooke J, Smith J M. Evolution of genetic redundancy. Nature, 1997, 388: 167−170
[25] Brown T A. Yuan J-G(袁建刚) trans. Genomes 2, 1st edn. Beijing: Beijing scientific Publishers, 2006. pp 538−544 (in Chinese)
[26] Barbazuk W B, Fu Y, McGinnis K M. Genome-wide analyses of alternative splicing in plants:opportunities and challenges. Genome Res,2008, 18: 1381−1392
[27] Zhang Z, Carriero N, Gerstein M. Comparative analysis of processed pseudogenes in the mouse and human genomes. Trends Genet, 2004, 20: 62−67
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