Cloning and Prokaryotic Expression Analysis of APX Gene from Maize

doi:10.7668/hbnxb.2014.04.009

Abstract

Abstract: Ascorbate peroxidase(APX)is a key enzyme which eliminates oxygen free radicals and increases plant resistance in adverse circumstances.In order to study the function of ascrobate peroxidose in cold treated maize leves.The complete open reading frame of APX gene was cloned from cold-stressed(4℃)maize inbred line E28 using the method of reverse transcription PCR(RT-PCR).The primers were designed according to the published APX cDNA sequences.The sequence of APX from E28 was 753 bp,encoding a protein of 250 amino acid residues with a predicted molecular weight of 27.3 kDa and a theoretical pI of 5.56.APX were constructed into the expression vector pET-28a(+)with full-length ORF,then transferred into E.coli.After inducing by IPTG,the product proteins of APX were detected by SDS-PAGE and the proteins were 27.3 kDa as expected.Salt tolerance analysis showed that the recombinant exhibit strong tolerance to salt than the non-recombinant bacteria,and the critical concentration of NaCl is 0.6 mol/L.Our researches could lay the foundation for the study of plant tolerance in the future.

Key words: APX, Prokaryotic expression, pET28a(+), IPTG, BL21(DE3)

CLC Number:

REN Ying, ZHAO Mei-ai, GUO Xin-mei, PEI Yu-he, SONG Xi-yun. Cloning and Prokaryotic Expression Analysis of APX Gene from Maize[J]. journal1, 2014, 29(4): 49-55. doi: 10.7668/hbnxb.2014.04.009.

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References

[1] Foyer C H,Halliwell B.The presence of glutathione and glutathione reductase in chloroplasts:A proposed role in ascorbic acid metabolism[J].Planta,1976,133(1):21-25.
[2] Nakano Y,Asada K.Hydrogen peroxide is scavenged by ascorbate-specific peroxidase in spinach chloroplasts[J].Plant Cell Physiology,1981,22(5):867-880.
[3] Sun W H,Mao W Q,Meng Q W.Functional mechanism and enzymatic and molecular characteristic of ascorbate peroxidase in plants[J].Plant Physiology Communications,2005,41(2):143-147.
[4] Amin M A.Role of dissolved oxygen reduction in improvement inhibition performance of ascorbic acid during copper corrosion in 0.50 mol/L sulphuric acid[J].Chinese Chemical Letters,2010,21(3):341-345.
[5] 战伟龑,张学杰,杨德翠,等.NaCl和等渗PEG对荞麦SOD及APX活性的影响[J].江苏农业科学,2009(5):101-102.
[6] 刘香娥,郭世荣,田婧,等.嫁接对西瓜幼苗生长及其耐盐性的影响[J].江苏农业学报,2009,25(3):628-634.
[7] 徐忠俊,胡凡,韩明明,等.镉胁迫对转基因水稻根和地上部抗氧化酶的影响[J].江苏农业科学,2009(6):92-94.
[8] 朴仁哲,赵洪颜,金玉姬,等.百草枯胁迫对地黄和苘麻膜脂过氧化及抗氧化酶活性的影响[J].江苏农业科学,2008(3):271-274.
[9] 马菁平,朱昌华,夏凯,等.卵磷脂和氯化胆碱对玉米幼苗生长及抗冷性的影响[J].江苏农业科学,2008(5):24-27.
[10] 张述义,刘玲玲.水分胁迫对不同品种小麦幼芽抗氧化酶活性的影响[J].山西农业科学,2013,41(2):122-125.
[11] 王文刚,李灵芝,张惠,等.镧对UV-C胁迫下番茄幼苗抗氧化酶活性的影响[J].山西农业科学,2011,39(6):532-534,542.
[12] 刘海,郭建文.高温胁迫对玉米生长期生理特性的影响[J].天津农业科学,2014,20(3):105-107.
[13] 张艳,林莺,刘永慧,等.NaCl对海滨锦葵活性氧清除能力的影响[J].山东师范大学学报:自然科学版,2007,22(4):117-119.
[14] 沈文飚,黄丽琴,徐朗莱.植物抗坏血酸过氧化物酶[J].生命的化学:中国生物化学会通讯,1997,7(5):26-28.
[15] 庞彩红,冯兰东,王宝山.高等植物中编码APX的基因特点及其与逆境响应的关系[J].山东师范大学学报:自然科学版,2007,22(4):114-116.
[16] Yoshimura K,Yabuta Y,Ishikawa T,et al.Expression of spinach ascorbate peroxidase isoenzymes in response to oxidative stresses[J].Plant Physiology,2000,123(1):223-234.
[17] Funatsuki H,Kurosaki H,Murakami T,et al.Deficiency of a cytosolic ascorbate peroxidase associated with chilling tolerance in soybean[J].Theor Appl Genet,2003,106(3):494-502.
[18] 段明.低温胁迫下番茄类囊体膜抗坏血酸过氧化氢酶基因的表达和功能研究[D].泰安:山东农业大学,2012.
[19] 孙卫红.番茄类囊体膜抗坏血酸过氧化物酶基因的克隆及功能分析[D].泰安:山东农业大学,2008.
[20] 张维静,陆海,杜希华.抗坏血酸过氧化物酶在植物抵抗氧化胁迫中的作用[J].山东师范大学学报:自然科学版,2008,23(4):113-115.
[21] 李泽琴,李静晓,张根发.植物抗坏血酸过氧化物酶的表达调控以及对非生物胁迫的耐受作用[J].遗传,2013,35(1):45-54.
[22] 孙卫红,李风,束德峰,等.转番茄正义抗坏血酸过氧化物酶基因提高烟草耐盐能力[J].中国农业科学,2009,42(4):1165-1171.

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