Scientia Agricultura Sinica ›› 2013, Vol. 46 ›› Issue (9): 1857-1866.doi: 10.3864/j.issn.0578-1752.2013.09.013

• HORTICULTURE • Previous Articles     Next Articles

Cloning, Sequence and Expression Analysis of NADP-Malic Enzyme Genes in Apple

 DONG  Qing-Long, WANG  Hai-Rong, AN  Miao, YU  Xian-Mei, WANG  Chang-Jun   

  1. Shandong Institute of Pomology, Tai’an 271000，Shandong
  • Received:2012-10-23 Online:2013-05-01 Published:2013-03-01

PDF

1048

Cited

20

Abstract: 【Objective】This study is aimed to characterize the genes of MdNADP-MEs involved in malic acid metabolism in Malus×domestica B. by gene cloning, sequence and expression of MdNADP-MEs in different tissues.【Method】 Three full-length cDNA sequences of MdNADP-ME1,2,3 were isolated by RT-PCR. The obtained cDNA sequences and the deduced amino acid sequences were analyzed with bioinformatics methods. qRT-PCR was used to assess the expression of MdNADP-MEs in different tissues.【Result】The sequencing results showed that three cDNAs (designated as MdNADP-ME1, 2 and 3; GenBank Accession No. JX971883, JX971884 and JX971885) were 1 512 bp, 1 782 bp and 1 926 bp. They contained an open reading frame (ORF) of 1 512 bp, 1 782 bp and 1 926 bp, respectively, and their ORFs encoded an protein with 503, 593 and 641 amino acids, respectively. Amino acid sequence and structure analysis indicated that MdNADP-MEs contained five conservative amino acid areas (motif I-V) and two functional structure domains: malic and NAD_bind_1_malic_enz. The result of phylogenetic analysis showed that MdNADP-ME1 and 2 belonged to cytosolic dicot NADP-ME (group I), MdNADP-ME3 belonged to plastidic dicot NADP-ME. qRT-PCR result showed that MdNADP-MEs were constitutively expressed in all examined tissues, but showed different expression levels.【Conclusion】MdNADP-ME1,2,3 belong to NADP-ME family, possess highly conserved structures, and show different expression patterns in different tissues of apple.

Key words: apple , malic enzyme , cloning , sequence analysis , expression analysis

[1]Detarsio E, Andreo C S, Drincocich M F. Basic residues play key roles in catalysis and NADP+-specificity in maize (Zea mays L.) photosynthetic NADP+-dependent malic enzyme. The Biochemical Journal, 2004, 382: 1025-1030.

[2]Bologna F P, Andreo C S, Drincovich M F. Escherichia coli malic enzymes: Two isoforms with substantial differences in kinetic properties metabolic regulation, and structure. Journal of Bacteriology, 2007, 189(16): 5937-5946.

[3]Wheeler M C G, Tronconi M A, Drincovich M F, Andreo C S, Flügge U, Maurino V G. A comprehensive analysis of the NADP-malic enzyme gene family of Arabidopsis. Plant Physiology, 2005, 139(1): 39-51.

[4]Drincovich M F, Casati P,  Andreo C S. NADP-malic enzyme from plants: a ubiquitous enzyme involved in different metabolic pathways. FEBS Letters, 2001, 490:1-6.

[5]刘畅, 李秀峰, 高野哲夫, 柳参奎. 高等植物中 NADP-ME的研究进展. 基因组学与应用生物学, 2011, 30: 1255-1260.

Liu C, Li X F, Tetsuo T, Liu S Q. Research progress of NADP-ME in higher plants. Genomics and Applied Biology, 2011, 30: 1255-1260. (in Chinese)

[6]Rothemel B A, Nelson T. Primary structure of maize NADP- dependent malic enzyme. Journal of Biology Chemistry, 1989, 264(33): 19587-19592. 

[7]Walter M H, Grima-Pettenati J, Grand C, Boudet A M, Lamb C J. Extensive sequence similarity of the bean CAD4 (cinnamyl-alcohol dehydrogenase) to a maize malic enzyme. Plant Molecular Biology, 1900,15(3): 525-526.

[8]Doorsseleare J V, Villaroel R, Montagu M V, Inze D. Nucleotide sequence of a cDNA encoding malic enzyme from poplar. Plant Physiology, 1991, 96(4): 1385-1386.

[9]Cushman J C. Characterization and expression of a NADP-malic enzyme cDNA induced by salt stress from the facultative crassulacean acid metabolism plant, Mesembryanthemum rystallinum. European Journal of Biochemistry, 1992, 208: 259-266.

[10]Franke K E, Adams D O. Cloning of a full-length cDNA for malic enzyme (EC 1.1.1.40) from grape berries. Plant Physiology, 1995, 107(3): 1009-1010. 

[11]Honda H, Shimada H, Akagi H. Isolation of a cDNA for an NADP-malic enzyme from Aloe arborescens. DNA Research, 1997, 4(6): 397-400.

[12]Honda H, Akagi H, Shimada H. An isozyme of the NADP-malic of a CAM plant, Aloe arborescens, with variation on conservative amino acid residues. Gene, 2000, 243(1/2): 85-92.

[13]Chi W, Yang J H, Wu N H, Zhang F. Four rice genes encoding NADP malic enzyme exhibit distinct expression profiles. Bioscience Biotechnology Biochemistry, 2004, 68(9): 1865-1874.

[14]Martinoia E, Rentsch D. Malate compartmentation- responses to a complex metabolism. Annual Review of Plant Physiology and Plant Molecular Biology, 1994, 45: 447- 467.

[15]Sutherland M W. The generation of oxygen radicals during host plant responses to infection. Physiological and Molecular Plant Pathology, 1991, 39: 79-93.

[16]Casati P, Drincovich M F, Edwards G E, Andreo C S. Malate metabolism by NADP-malic enzyme in plant defense. Photosynthesis Research, 1999, 61(2): 99-105.

[17]Fu Z Y, Zhang Z B, Hu X J, Shao H B, Ping X. Cloning, identification, expression analysis and phylogenetic relevance of two NADP- dependent malic enzyme genes from hexaploid wheat. Comptes Rendus Biologies, 2009, 332: 591-602.

[18]Goodenough P W, Prosser I M, Young K. NADP-linked malic enzyme sand malatemetabolism in ageing tomato fruit. Phytochemistry, 1985, 24(6): 1157-1162.

[19]Yao Y X, Li M, Liu Z, Hao Y J, Zhai H. A novel gene, screened by cDNA-AFLP approach, contributes to lowering the acidity of fruit in apple. Plant Physiology and Biochemistry, 2007, 45: 139-145.

[20]Dong Q L, Liu D D, An X H, Hu D G, Yao Y X, Hao Y J. MdVHP1 encodes an apple vacuolar H+-PPase and enhances stress tolerance in transgenic apple callus and tomato. Journal of Plant Physiology, 2011, 168: 2124-2133.

[21]Livak K J, Schmittgen T D. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods, 2001, 25: 402-408.

[22]Yang Z, Zhang H, Huang H, Kuo C, Tsai L, Yuan H S, Chou W, Chang G, Tong L. Structural studies of the pigeon cytosolic nadp+ -dependent malic enzyme. Protein Science, 2002, 11: 332-341.

[23]Yao Y X, Li M, Liu Z, You C X, Wang D M, Zhai H, Hao Y J. Molecular cloning of three malic acid related genes MdPEPC, MdVHA-A, MdcyME and their expression analysis in apple fruits. Scientia Horticulturae, 2009, 122: 404-408.

[24]Wheeler M C G, Arias C L, Tronconi M A, Maurino V G, Andreo C S, Drincovich M F. Arabidopsis thaliana NADP-malic enzyme isoforms: high degree of identity but clearly distinct properties. Plant Molecular Biology, 2008, 67(3): 231-242.

[25]Pengelly J J L, Tan J, Furbank R T, Caemmerer S. Antisense reduction of NADP-malic enzyme in Flaveria bidentis reduces flow of CO2 through the C4 cycle. Plant Physiology, 2012, 112: 203-240.

[26]Muller G L, Drincovich M F, Andreo C S, Lara M V. Nicotiana tabacum NADP-Malic enzyme: cloning, characterization and analysis of biological role. Plant Cell Physiology, 2008, 49(3): 469-480.
[1] SU YiFan, YANG ZhanXu, WANG Di, MAO JunCheng, WEI MengMeng, CHEN Ze, BAI XinRan, CHU TianGe, MA ChangNing, QIAO MingFei, SUN Quan, HU DaGang. Effects of 2, 4-Epibrassinolide on Postharvest Storage Quality and Physiological Performance of Apple [J]. Scientia Agricultura Sinica, 2026, 59(7): 1536-1551.
[2] YANG CaiLi, LI YongZhou, HE LiangLiang, SONG YinHua, ZHANG Peng, LIU ZhaoXian, LI PengHui, LIU SanJun. Genome-Wide Identification and Analysis of TPS Gene Family and Functional Verification of VvTPS4 in the Formation of Monoterpenes in Grape [J]. Scientia Agricultura Sinica, 2025, 58(7): 1397-1417.
[3] TENG MengXin, XU Ya, HE Jing, WANG Qi, QIAO Fei, LI JingYang, LI XinGuo. Identification and Functional Analysis of Ca2+-ATPase Gene Family in Banana [J]. Scientia Agricultura Sinica, 2025, 58(7): 1418-1433.
[4] ZHENG YaQin, LIU XueQing, WU SiWen, TANG XiaoYan, YANG DanNi, WANG YongKang, AHMAD Aftab, KHAN Afrsyab, WANG ChengGang, CHEN GuoHu. Cloning and Expression of BcDET2 Gene and Functional of Its Regulatory Effect on Bolting and Flowering in Wucai (Brassica campestris L.) [J]. Scientia Agricultura Sinica, 2025, 58(5): 991-1003.
[5] CONG QiQi, ZHANG JingYi, MENG XiangLong, DAI PengBo, LI Bo, HU TongLe, WANG ShuTong, CAO KeQiang, WANG YaNan. Identification of Hypovirus in Apple Ring Rot Fungus Botryosphaeria dothidea and Detection of Virus-Carrying Status in China [J]. Scientia Agricultura Sinica, 2025, 58(3): 478-492.
[6] ZHANG LinLin, GONG Rui, CUI YanLing, ZHONG XiongHui, LI Ye, LI RanHong, QIAN ZongWei. Effect Analysis of SmWRKY30 in Eggplant Resistance to Ralstonia solanacearum by Virus Induced Gene Silencing (VIGS) [J]. Scientia Agricultura Sinica, 2025, 58(3): 548-563.
[7] PAN Yuan, WANG De, LIU Nan, MENG XiangLong, DAI PengBo, LI Bo, HU TongLe, WANG ShuTong, CAO KeQiang, WANG YaNan. Evaluation of the Effectiveness of Two High-Throughput Sequencing Techniques in Identifying Apple Viruses and Identification of Two Novel Viruses [J]. Scientia Agricultura Sinica, 2025, 58(2): 266-280.
[8] ZOU PeiYi, LIU MeiYan, WANG Ying, LI RanHong. Cloning and Functional Study of AkNAC2 from Actinidia kolomikta [J]. Scientia Agricultura Sinica, 2025, 58(19): 3985-3999.
[9] YI ZeHui, WANG Ying, SONG HuiXia, ZHAO Jing, MAO LiPing. Genome-Wide Identification and Expression Analysis of Peroxiredoxins Gene Family in Asparagus officinalis [J]. Scientia Agricultura Sinica, 2025, 58(18): 3728-3743.
[10] ZHANG ShuHong, GAO FengJu, WU QiuYing, JI JingXin, ZHANG YunFeng, XU Ke, GU ShouQin, FAN YongShan. Cloning and Expression Analysis of Heat Shock Protein HSP 9/12 Genes in Setosphaeria turcica [J]. Scientia Agricultura Sinica, 2025, 58(18): 3648-3663.
[11] QI XiangYu, LI XinRu, CHEN ShuangShuang, FENG Jing, CHEN HuiJie, LIU XinTong, JIN YuYan, DENG YanMing. Identification of the FLA Gene Family and Functional Analysis of JsFLA2 in Jasminum sambac [J]. Scientia Agricultura Sinica, 2025, 58(17): 3516-3530.
[12] ZENG YueHui, ZOU WenGuang, ZHAO FuMing, XIAO ChangChun, HUANG JianHong, MA BinLin, YANG WangXing, WEI XinYu, XU XuMing. Map-Based Cloning and Functional Verification of A Novel Split Glume Gene OsSG2 in Rice (Oryza sativa L.) [J]. Scientia Agricultura Sinica, 2025, 58(11): 2062-2080.
[13] LÜ ShuWei, TANG Xuan, LI Chen. Research Progress on Seed Shattering of Rice [J]. Scientia Agricultura Sinica, 2025, 58(1): 1-9.
[14] WANG Wei, WU ChuanLei, HU XiaoYu, LI JiaJia, BAI PengYu, WANG GuoJi, MIAO Long, WANG XiaoBo. Genome-Wide Identification of Soybean LOX Gene Family and the Effect of GmLOX15A1 Gene Allele on 100-Seed Weight [J]. Scientia Agricultura Sinica, 2025, 58(1): 10-29.
[15] GUAN ZhiLin, JIN FengWei, LIU TingTing, WANG Yi, TAN YingYing, YANG ChunHui, LI RuiTong, WANG Bo, LIU KeDe, DONG Yun. Genetic Analysis and Gene Mapping of Glossy Leaf in Brassica napus [J]. Scientia Agricultura Sinica, 2024, 57(4): 650-662.
Viewed
Full text


Abstract

Cited
  Shared   
  Discussed   
No Suggested Reading articles found!
京ICP备09089781号-30
Copyright 2018 © Editorial office of Scientia Agricultura Sinica
Tel: 86-010-82106279    E-mail: zgnykx@mail.caas.net.cn
Supported by Beijing Magtech Co.Ltd