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作物学报 ›› 2011, Vol. 37 ›› Issue (07): 1134-1143.doi: 10.3724/SP.J.1006.2011.01134

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

小麦生理型和遗传型雄性不育系及其保持系小花完整叶绿体蛋白质组分比较研究

李莉,王书平**,张改生*,王亮明,宋瑜龙,张龙雨,牛娜,马守才   

  1. 西北农林科技大学陕西省作物杂种优势研究与利用重点实验室 / 小麦育种教育部工程研究中心,陕西杨凌 712100?
  • 收稿日期:2010-12-23 修回日期:2011-03-27 出版日期:2011-07-12 网络出版日期:2011-05-11
  • 通讯作者: 张改生, E-mail: zhanggsh@public.xa.sn.cn, Tel: 029-87092152
  • 基金资助:

    本研究由国家高技术研究发展计划(863计划)重大专项(2009AA101102), 国家自然科学基金项目(31071477), 高等学校博士学科点专项科研基金(20090204110024), 陕西省“13115”科技创新工程重大科技专项(2010ZDKG-68)和西北农林科技大学拔尖人才支持计划项目资助。

Comparison of Floret Intact Chloroplast Proteome in Male Sterile Line induced by CHA-SQ-1, Cytoplasmic-Nuclear Sterile and Its Normal Fertile Lines in Wheat

LI Li,WANG Shu-Ping**,ZHANG Gai-Sheng*,WANG Liang-Ming,SONG Yu-Long,ZHANG Long-Yu,NIU Na,MA Shou-Cai   

  1. Key Laboratory of Crop Heterosis of Shaanxi Province, Northwest A&F University / Wheat Breeding Engineering Research Center, Ministry of Education, Yangling 712100, China
  • Received:2010-12-23 Revised:2011-03-27 Published:2011-07-12 Published online:2011-05-11
  • Contact: 张改生, E-mail: zhanggsh@public.xa.sn.cn, Tel: 029-87092152

摘要: 采用固相pH梯度SDS聚丙烯酰胺双向凝胶电泳技术,以小麦遗传型雄性不育系ms(S)-1376、杀雄剂SQ-1诱导的生理型雄性不育系ms(A)-1376,以及对应的保持系(A)-1376 (杀雄剂诱导前的正常可育系)所构建的等基因和等生理差异系为试材,比较分析了2种分离纯化完整叶绿体的方法。在确立了一套适用的技术方法的基础上,研究了三者在花粉小孢子萌发单核早期小花完整叶绿体蛋白之间的差异。采用30%、45%和60%的不连续蔗糖密度梯度离心的方法能够获得完整且纯度较高的叶绿体,经TCA-丙酮提取蛋白质后,PDQuest软件分析,在分子量14.4~66.2 kD,等电点4~7的线性范围内可分辨出2-DE图谱上239个较为清晰的蛋白质点,对其中的6个差异表达蛋白质点进行MALDI-TOF肽质量指纹图谱分析,从生物信息学数据库检索鉴定出6个差异蛋白质点分别是酰基辅酶A脱氢酶结构域蛋白、钙调结合蛋白磷酸酶、多催化功能肽酶、热休克蛋白60、光受体蛋白2及1个未知功能的表达蛋白质。这些蛋白质在供试小麦叶绿体物质能量代谢、叶绿体防卫抵御机制、叶绿体细胞内信号转导及植物极性生长等生理应答反应中起调控作用,它们能在本研究供试两不育系和对应可育系中差异表达,可能与雄性不育相关。

关键词: 小麦, 遗传型和生理型雄性不育, 完整叶绿体蛋白质组, 差异表达, 2D-PAGE, MALDI-TOF-MS

Abstract: Weextracted floret chloroplast proteins from the genetic male sterile line ms(S)-1376, the physiological male sterile line ms(A)-1376 induced by chemical hybridizing agent SQ-1, and their counterpart maintainer line (A)-1376 (normal fertility). These proteins were separated via 2-D electrophoresis with immobilized pH 4–7 gradients as the first dimension and SDS-PAGE as the second one. We established a set of technique available for intact chloroplast isolation and 2-DE of proteins in wheat floret and compared the effect of intact chloroplast purification using two methods. Based on the 2-DE technique, we explored the difference of the floret chloroplast proteins among the three materials tested in their early uninucleate anther stage. The result showed that we obtained high purity of intact chloroplast using discontinuous sucrose density gradient centrifugation with three-step gradient densities of 30%, 45%, and 60%. Proteins were extracted from chloroplast by TCA-acetone and analyzed by PDQuest software, we got as much as 239 protein spots within pH 4–7 and with molecular weight of 14.4–66.2 kD on each 2-DE gel. We analyzed six differential expressed proteins by matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry (MS) and bioinformatical database, and found they were acyl-CoA dehydrogenase domain protein, calmodulin-binding protein phosphatase, multiple catalytic peptidase, heat-shock protein 60, light receptor protein 2 and a protein of unknown function. These proteins are involved in series of physiological reactions such as metabolism of energy substances, chloroplast defendance, chloroplasts signal transduction and plant growth, the differential expression of those proteins among the three materials tested is likely related to the male sterility in wheat.

Key words: Wheat, Genetic and physiological male sterilities, Intact chloroplast proteins, Differential expression, 2-dimensional electrophoresis, MALDI-TOF-MS

[1]Liu Z-C(刘祚昌), Li J-G(李继耕), Luo H-X(罗会馨), Chen F-T(陈福太). Studies on the ribulose bisphosphate carboxylase and cytoplasmic male-sterility. Acta Genet Sin (遗传学报), 1983, 10(1): 36-42 (in Chinese with English abstract)
[2]Pei Y-X(裴雁曦), Chen Z-J(陈竹君), Cao J-S(曹家树). Comparative study on polypeptides of mitochondria and chloroplast between the male cytoplasmic sterile line and its maintainer line in tuber mustard (Brassica juncea Coss. var. tumida). Acta Bot Boreali-Occident Sin (西北植物学报), 2004, 24(8): 1511-1513 (in Chinese with English abstract)
[3]Li J-G(李继耕), Gao W-Q(高文琴). Studies on the chloroplast ultrastructure in the cytoplasmic male-sterile line and its maintainer of Brassica napus. Acta Genet Sin (遗传学报), 1983, 10(4): 280-282 (in Chinese with English abstract)
[4]Li J-Y(李家洋), Li J-G(李继耕). Chloroplast thylakoid membrane polypeptides and cytoplasmic male sterility. Acta Genet Sin (遗传学报), 1986, 13(6): 430-436 (in Chinese with English abstract)
[5]Ying Y-R(应燕如), Ni D-Z(倪大洲), Cai Y-X(蔡以欣). Comparative analysis of the fraction i protein in rice, wheat, rape and tobacco cytoplasmic male sterile systems. Acta Genet Sin (遗传学报), 1989, 16(5): 362-366 (in Chinese with English abstract)
[6]Zhang L-Y(张龙雨), Li H-X(李红霞), Zhang G-S(张改生), Wang J-S(王俊生), Han Y-F(韩艳芬), Yuan Z-J(袁正杰), Niu N(牛娜), Ma S-C(马守才). Cloning and expression analysis of cMDH gene related to cytoplasmic male sterile wheat with Aegilops kotschyi cytoplasm. Acta Agron Sin (作物学报), 2009, 35(9): 1620-1627 (in Chinese with English abstract)
[7]Guo Y-P(郭艳萍), Wei F(位芳), Zhang G-S(张改生), Cheng H-G(程海刚), Song Y-L(宋瑜龙), Wang Q(王青), Niu N(牛娜), Ma S-C(马守才), Li H-X(李红霞). Molecular and cytological analyses of 1BL/1RS wheat and fertility-restoring locations for male sterile lines with Aegilops cytoplasm. Sci Agric Sin (中国农业科学), 2010, 43(14): 2839-2847 (in Chinese with English abstract)
[8]Wang J-S(王俊生), Yuan L(袁蕾), Zhang M-Z(张明珠), Zhang G-S(张改生), Niu N(牛娜), Ma S-C(马守才), Li H-X(李红霞). Expression of Ubiquitin-26S proteasome pathway in physiological male sterility of wheat induced by chemical hybrid agents SQ-1. J Agric Biotechnol (农业生物技术学报), 2010, 18(4): 695-701 (in Chinese with English abstract)
[9]Zhang M-Z(张明珠), Yuan L(袁蕾), Wang J-S(王俊生), Zhang G-S(张改生), Guo W-J(郭文江), Zhang L-Y(张龙雨). Expressing analysis of cACO gene in Male sterile wheat induced by chemical hybridizing agent SQ-1. Chin J Biochem Mol Biol (中国生物化学与分子生物学报), 2010, 26(8): 740-748 (in Chinese with English abstract)
[10]Elias B A, Givan C V. Density gradient and differential centrifugation methods for chloroplast purification and enzyme localization in leaf tissue. Planta, 1978, 142: 317-320
[11]Chen Y-W(陈云伟), Zhang N-H(张年辉), Zhao Y(赵云), Du L-F(杜林方), Nakai M. Preparation of envelope membrane of rape chloroplast and detection of Toc33. Plant Physiol Commun (植物生理学通讯), 2004, 40(2): 223-225 (in Chinese)
[12]Sun Q(孙群), Hu J-J(胡景江). Techniques for Plant Physiology Research (植物生理学研究技术). Yangling: Northwest A&F University Press, 2005. pp 127-131 (in Chinese)
[13]Chen R-H(陈蕊红), Ye J-X(叶景秀), Zhang G-S(张改生), Wang J-S(王俊生), Niu N(牛娜), Ma S-C(马守才), Zhao J-X(赵继新), Zhu J-C(朱建楚). Differential proteomic analysis of anther proteins between cytoplasmic-nuclear male sterility line and its maintainer in wheat (Triticum aestivum L.). Prog Biochem Biophys (生物化学与生物物理进展), 2009, 36(4): 431-440 (in Chinese with English abstract)
[14]Bradford M M. A rapid and sensitive method for the quantitation of microgram quantities utilizing the principle of protein dye binding. Anal Biochem, 1976, 72: 248-254
[15]Ye J-X(叶景秀), Chen R-H(陈蕊红), Zhang G-S(张改生), Wang J-S(王俊生), Wang S-P(王书平), Li L(李莉), Niu N(牛娜), Ma S-C(马守才), Li H-X(李红霞), Zhu J-C(朱建楚). Analysis on anther proteins of wheat male sterile line induced by chemical hybridizing agent SQ-1. J Agric Biotechnol (农业生物技术学报), 2009, 17(5): 858-864 (in Chinese with English abstract)
[16]Ye J-X(叶景秀), Zhang G-S(张改生), Wang S-P(王书平), Chen R-H(陈蕊红), Wang J-S(王俊生), Niu N(牛娜), Ma S-C(马守才), Li H-X(李红霞), Zhu J-C(朱建楚). Differential proteomic studies on pollen grain proteins of wheat male sterile line induced by chemical hybridizing agent SQ. Chin J Biochem Mol Biol (中国生物化学与分子生物学报), 2009, 25(10): 949-957 (in Chinese with English abstract)
[17]Zhao H-J(赵虎基), Wang J-H(王建华), Gao P(高鹏), Gu R-L(顾日良), Zhang J-Q(张京强), Wang T-Y(王天宇), Wang G-Y(王国英). Cloning of plastid acetyl-CoA carboxylase cDNA from setariaitalica and sequence analysis of graminicide target site. Acta Bot Sin (植物学报), 2004, 46(6): 751-756 (in Chinese with English abstract)
[18]Liu S-H(刘诗航), Wang C-X(王彩香), Mao X-G(毛新国), Liu H-M(刘惠民), Li A(李昂), Jing R-L(景蕊莲). Cloning of protein phosphatase 2A regulatory subunit gene TaBβ-1 and its expression patterns under abiotic stresses in wheat. Sci Agric Sin (中国农业科学), 2010, 43(11): 2197-2208 (in Chinese with English abstract)
[19]Tang Z(唐征), Yang K(杨凯), Feng Y-Q(冯永庆), Qin L(秦岭). Cloning and homology analysis of chestnut protein phosphatase PP2A catalytic subunit gene. J Beijing Univ of Agric(北京农学院学报), 2009, 24(2): 1-4 (in Chinese with English abstract)
[20]Ma B-J(马伯军), Gu Z-M(顾志敏), Tang H-J(唐海娟), Chen X-F(陈析丰), Liu F(刘峰), Zhang H-S(张红生). Preliminary study on the function of calcineurin B-like protein gene OsCBL8 in rice. Chin J Rice Sci (中国水稻科学), 2009, 23(2): 127-134 (in Chinese with English abstract)
[21]Kong L A, Wang M, Wang Q L, Wang X H, Lin J X. Protein phosphatase 1 and /or 2A is involved in the regulation of calcium uptake and pollen tube development in Picea wislonii. Tree Physiol, 2006, 26: 1001-1012
[22]Xiang X(向珣), Li C-Q(李成琼), Lei J-J(雷建军). Research of HSPs and heat resistance. Northern Hortic (北方园艺), 1999, (4): 9-10 (in Chinese)
[23]Zhang J-H(张俊环), Huang W-D(黄卫东). Cold acclimation induce the changes in the sub-cellular distribution of small heat shock protein 17.6 in young grape leaves during heat stress. Acta Agric Boreali-Sin (华北农学报), 2009, 24(2): 8-11 (in Chinese with English abstract)
[24]Wang L, Zhao C M, Wang Y J, Liu J. Over-expression of chloroplast-localized small molecular heat-shock protein enhances chilling tolerance in tomato plant. Acta Photophysiol Sin, 2005, 31(2): 167-174
[25]Yang C-Y(杨传燕), Wang C(王翠), Zhang J-X(张景霞), Wang J-Y(王佳颖), Wang L(王丽), Liu J(刘箭). Over-expression of chloroplast- localized small molecular heat-shock protein enhances the thermotolerance in plant. J Shandong Normal Univ (Nat Sci) (山东师范大学学报•自然科学版), 2008, 23(4): 106-108 (in Chinese with English abstract)
[26]Stoelzle S, Kagawa V, Wada M, Hedrich R, Dietrich P. Blue light activates calcium-permeable channels in Arabidopsis mesophyll cells via the phototropin signaling pathway. Proc Natl Acad Sci USA, 2003, 100: 1456-1461
[27]Gilliberto L, Perrotta G, Pallara P. Manipulation of the blue light photo-receptor cryptochrome 2 intomato affects vegetative development, flower in time, and fruit antioxidant content. J Plant Physiol, 2005, 137: 199-208
[28]Cao S-H(曹双河), Zhang X-Q(张相岐), Zhang A-M(张爱民). Review of the molecular regulation mechanism and genetics of photoperiod-and/or thermosensitive male sterility. Bull Bot (植物学通报), 2005, 22(1): 19-26 (in Chinese with English abstract)
[29]Cui Y-L(崔永兰), Wang P-C(王鹏程), Yang Z-N(杨仲南), Zhang S(张森). Chloroplast localization of the expressed protein encoded by At3g61870 in Arabidopsis thaliana. Acta Bot Sin (西北植物学报), 2008, 28(4): 662-666 (in Chinese with English abstract)
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