|本期目录/Table of Contents|

[1]赖先军,古英洪,王海燕,等.甘薯Sporamin家族基因克隆与表达分析[J].应用与环境生物学报,2013,19(02):215-223.[doi:10.3724/SP.J.1145.2013.00215]
 LAI Xianjun,GU Yinghong,WANG Haiyan,et al.Cloning and Expression Analyses of Sporamin Family Genes in Sweet Potato[J].Chinese Journal of Applied & Environmental Biology,2013,19(02):215-223.[doi:10.3724/SP.J.1145.2013.00215]
点击复制

甘薯Sporamin家族基因克隆与表达分析()
分享到:

《应用与环境生物学报》[ISSN:1006-687X/CN:51-1482/Q]

卷:
19卷
期数:
2013年02期
页码:
215-223
栏目:
研究论文
出版日期:
2013-04-25

文章信息/Info

Title:
Cloning and Expression Analyses of Sporamin Family Genes in Sweet Potato
作者:
赖先军古英洪王海燕张义正
(四川大学生命科学学院,四川省分子生物学及生物技术重点实验室 成都 610064)
Author(s):
LAI Xianjun GU Yinghong WANG Haiyan ZHANG Yizheng
(Sichuan Key Laboratory of Molecular Biology & Biotechnology, College of Life Sciences, Sichuan University, Chengdu 610064, China)
关键词:
甘薯储藏蛋白基因克隆序列分析数字基因表达谱蛋白酶抑制剂
Keywords:
sweet potato sporamin gene cloning sequence analysis digital gene expression profiling protease inhibitor
分类号:
Q946.1 : Q78
DOI:
10.3724/SP.J.1145.2013.00215
文献标志码:
A
摘要:
为研究甘薯sporamin家族基因的结构和功能,通过对本实验室构建的甘薯转录组序列进行分析,设计相应引物并采用高保真DNA聚合酶进行PCR扩增,经TA克隆和序列测定,利用在线软件对甘薯sporamin结构进行分析和功能预测. 从转录组数据库中共发现61条sporamin contigs,其中sporamin A家族有31条,sporamin B家族有30条,已从甘薯中克隆到6个sporamin A基因和10个sporamin B基因. 所有sporamin A基因编码区全长660 bp,基因间序列一致性为95%以上;sporamin B基因编码区全长651 bp,基因间序列一致性达到98%;两个基因家族间序列一致性为84%. 对克隆的sporamin A和sporamin B进行的数字表达谱分析结果表明,sporamin A和B在膨大初期的块根和成熟块根中的转录水平最高,在膨大中期转录水平较低,在叶片和茎中表达量极低. Sporamin氨基酸保守序列和同源比对结果显示,该蛋白质含有STI(Soybean Trypsin Inhibitor)保守序列,此序列有170多个氨基酸,属于Kunitz型蛋白酶抑制剂超家族,表明该蛋白具有蛋白酶抑制剂功能. 图9 表3 参33
Abstract:
In order to study the structure and function of sporamin family genes in sweet potato, the transcriptome database of this plant constructed in our lab were analyzed to screen sporamin contigs and appropriate primers designed to amplify sporamin genes, which were then cloned and sequenced. All together 61 sporamin contigs were found in the transcriptome database, including 31 contigs in sporamin A sub-family and 30 contigs in sporamin B sub-family. Six sporamin A genes and 10 sporamin B genes were cloned and sequenced. ORF of all sporamin A genes were 660 bp in length with identity more than 95% among all sporamin A genes. ORF of sporamin B genes had a total length of 651 bp with identity more than 98% among sporamin B genes. Homology between the two sub-families was 84%. The results of digital gene expression profile analysis showed that both sporamin A and B had the highest level of transcription in the initial tuberous roots and harvest tuberous roots, comparatively low transcription in the expanding tuberous roots, and very little in leaves and stems. Analysis of conserved sequence of sporamin genes and homology comparison displayed that the proteins contained a soybean trypsin inhibitor conserved sequence with more than 170 amino acids belonging to the Kunitz-type protease inhibitor superfamily, indicating that the sporamin has the function of protease inhibitors. Fig 9, Tab 3, Ref 33

参考文献/References:

1 Maeshima M, Sasaki T, Asahi T. Characterization of majorproteins in sweet potato tuberous roots [J]. Phytochemistry, 1985, 24: 1899-1902
2 Hattori T, Nakamura K, Yoshida. Structural relationship among the members of a multi-gene family coding for the sweet potato tuberous root storage protein [J]. Plant Mol Biol, 1989, 13: 563-572
3 Murakami S, Hattori T, Nakamura K. Structural differences in full-length cDNAs for two classes of sporamin, the major soluble protein of sweet potato tuberous roots [J]. Plant Mol Biol, 1986, 7: 343-355
4 Wettstein D, Chua NH. Plant Molecular Biology [M]. New York: Plenum Press, 1987
5 Conlan RS, Griffiths LA, Napier JA, Shewry PR, Mantell S, Ainsworth C. Isolation and characterisation of cDNA clones representing the genes encoding the major tuber storage protein (dioscorin) of yam (Dioscorea cayenensis Lam.) [J]. Plant Mol Biol, 1995, 28 (3): 369-380
6 Yang J, Barr LA, Fahnestock SR, Liu ZB. High yield recombinant silk-like protein production in transgenic plants through protein targeting [J]. Transgenic Res, 2005, 14: 313-324
7 Woofe JA. Sweet Potato, An Untapped Food Resource [M]. New York: Cambridge University Press, 1992
8 Yeh KW, Chen JC, Lin MI, Chen YM, Lin CY. Functional activity of sporamin from sweet potato (Ipomoea batatas Lam.): a tuber storage protein with trypsin inhibitory activity [J]. Plant Mol Biol, 1997, 33: 565-570
9 Yeh KW, L in MI, Tuan SJ, Chen YM, Lin CJ, Kao SS. Sweet potato (Ipomoea batatas) trypsin inhibitors expressed in transgenic tobacco plants confer resistance against Spodoptera litura [J]. Plant Cell Rep, 1997, 16 (10): 696-699
10 Koiwa H, Shade RE, Zhu-Salzman K, D’Urzo MP, Murdock LL, Bressan RA, Hasegawa PM. A plant defensive cystatin (soyacystatin) targets cathepsin L-Like digestive cysteine proteinases (DvCALs) in the larval midgut of western corn rootworm (Diabrotica virgifera) [J]. FEBS Lett, 2000, 471: 67-70
11 Hong YF, Liu CY, Cheng KJ, Hour AL, Chan MT, Tseng TH, Chen KY, Shaw JF, Yu SM. The sweet potato sporamin promoter confers high-level phytase expression and improves organic phosphorus acquisition and tuber yield of transgenic potato [J]. Plant Mol Biol, 2008, 67: 347-361
12 邱琳, 董衡, 黄鹂, 曹家树. 甘薯sporamin蛋白的功能及其在抗虫转基因植物中的应用[J]. 中国细胞生物学学报, 2011, 33 (6): 692-698 [Qiu L, Dong H, Huang L, Cao JS. The function of sweet potato’s special tuber storage protein and its application in engineering plant research [J]. Chin J Cell Biol, 2011, 33 (6): 692-698]
13 Hurley BA, Tran HT, Marty NJ, Park J, Snedden WA, Mullen RT, Plaxton WC. The dual-targeted purple acid phosphatase isozyme At PAP26 is essential for efficient acclimation of Arabidopsis to nutritional phosphate deprivation [J]. Plant Physiol, 2010, 7 (153): 1112-22
14 陈克贵, 王海燕, 张义正. 甘薯贮藏蛋白(sporamin) A基因编码区克隆及序列同源性分析[J]. 中国生物化学与分子生物学报, 2000, 16 (1): 41-45 [Chen KG, Wang HY, Zhang YZ. Cloning and homologous analysis of sporamin A gene coding region sequence of Ipomoea batatas [J]. Chin J Biochem Mol Biol, 2000, 16 (1): 41-45]
15 封树梅, 张义正. 甘薯储藏蛋白(sporamin) A基因在大肠杆菌中的表达研究[J]. 四川大学学报(自然科学版), 2000, 37 (3): 446-450 [Feng SM, Zhang YZ. Expression of sporamin A gene of sweet potato in Escherichia coli [J]. Jf Sichuan Univ (Nat Sci Ed), 2000, 37 (3): 446-450]
16 Tao X, Gu YH, Wang HY, Zheng W, Li X, Zhao CW, Zhang YZ. Digital gene expression analysis based on integrated de novo transcriptome assembly of sweet potato [Ipomoea batatas (L.) Lam.] [J]. PLoS ONE, 2012, 7 (4): e36234
17 Sambrook J, Fritsch EF, Maniatis T. Molecular Cloning: A Laboratory Manual [M]. 2nd ed. New York: Cold Spring Harbor Laboratory Press, 1989
18 Loebenstein G. Origin, distribution and economic importance [M]. The Sweetpotato. Springer Netherlands, 2009: 9-12
19 Trapnell C, Pachter L, Salzberg SL. Discovering splice junctions with RNA-Seq [J]. Bioinformatics, 2009, 25 (9): 1105-1111
20 程龙军, 郭得平, 葛红娟. 甘薯块根特异蛋白——sporamin的研究进展[J]. 植物学通报, 2001, 18 (6): 672-677 [Cheng LJ, Guo DP, Ge HJ. The special proteins in sweet potato tuber-sporamin [J]. Chin Bull Bot, 2001, 18 (6): 672-677]
21 Hattori T, Nakamura K. Genes coding for the major tuberous root protein of sweet potato: identification of putative regulartory sequence in the 5’ upstream region [J]. Plant Mol Biol, 1988, 22: 417-426
22 Chen JC, Chen YM, Yeh KW. Isolation and characterization of new sporamin gene members from sweet potato (Imomoea batatas Lam.) [J]. Taiwania, 1997, 42: 34-42
23 Ohta S, Hattori T, Morikami A. High-level expression of a sweet potato sporamin gene promoter β-glucuroidase (GUS) fusion gene in the stems of transgenic tobacco plants is conferred by multiple cell type-specific regulatory elements [J]. Mol Gen Genet, 1991, 225: 369-378
24 Marchler-Bauer A, Lu S, Anderson JB, Chitsaz F, Derbyshire MK, Weese-Scott CD, Fong JH, Geer LY, Geer RC, Gonzales NR, Gwadz M, Hurwitz DI, Jackson JD, Ke Z, Lanczycki CJ, Lu F, Marchler GH, Mullokandov M, Omelchenko MV, Robertson CL, Song JS, Thanki N, Yamashita RA, Zhang D, Zhang N, Zheng C, Bryant SH. CDD: a Conserved Domain Database for the functional annotation of proteins [J]. Nucleic Acids Res, 2011, 39D: 225-229
25 Hattori T, Nakagawa T, Maeshima M. Molecular cloning and nucleotide sequence of cDNA for sporamin, the major soluble protein of sweet potato tuberous roots [J]. Plant Mol Biol, 1985, 5: 313-320
26 Nakamura K, Matsushita K, Mukumoto F. Precursor to sweet potato sporamin in transformed tobacco cell line BY-2 [J]. J Exp Bot, 1993, 44: 331-338
27 Matsuoka K, Matsuomoto S, Hattori T. Vacuolar targeting and post-translational processing of the precursor to the sweet potato tuberous root storage protein in heterologous plant cells [J]. J Biol Chem, 1990, 265: 19750-19757
28 Arnold K, Bordoli L, Kopp J, Schwede T. A web-based environment for protein structure homology modelling [J]. Bioinformatics, 2006, 22: 195-201
29 Schwede T, Kopp J, Guex N, Peitsch MC. An automated protein homology-modeling server [J]. Nucleic Acids Res, 2003, 31: 3381-3385
30 Guex N, Peitsch MC. An environment for comparative protein modelling [J]. Electrophoresis, 1997, 18: 2714-2723
31 Yao PL, Hwang MJ, Chen YM, Yeh KW. Site-directed mutagenesis evidence for a negatively charged trypsin inhibitory loop in sweet potato sporamin [J]. FEBS Lett, 2001, 496: 134-138
32 Huang GJ, Sheu MJ, Chen HJ, Chang YS, Lin YH. Growth inhibition and induction of apoptosis in NB4 promyelocytic leukemia cells by trypsin inhibitor from sweet potato storage roots [J]. J Agric Food Chem, 2007, 55: 2548-2553
33 Yao J, Qian CJ. Sporamin induce apoptosis in human tongue carcinoma cells by down-regulating Akt/GSK-3 signaling [J]. Fundam Clin Pharmacol, 2011, 25 (2): 229-236

相似文献/References:

[1]靳艳玲,甘明哲,周玲玲,等.4个甘薯品种不同生育期的乙醇发酵比较[J].应用与环境生物学报,2009,15(02):262.[doi:10.3724/SP.J.1145.2009.00267]
 JIN Yanling,GAN Mingzhe,et al.Ethanol Production with 4 Varieties of Sweet Potato at Different Growth Stages[J].Chinese Journal of Applied & Environmental Biology,2009,15(02):262.[doi:10.3724/SP.J.1145.2009.00267]
[2]陶向,张勇为,姜玉松,等.甘薯块根储藏过程中的淀粉含量变化[J].应用与环境生物学报,2010,16(05):741.[doi:10.3724/SP.J.1145.2010.00741]
 TAO Xiang,ZHANG Yongwei,JIANG Yusong,et al.Changes in Root Starch Contents of Sweet Potato Cultivars During Storage[J].Chinese Journal of Applied & Environmental Biology,2010,16(02):741.[doi:10.3724/SP.J.1145.2010.00741]
[3]杨俊仕,周后珍,李国欣,等.组合工艺处理甘薯燃料乙醇糟液[J].应用与环境生物学报,2010,16(05):730.[doi:10.3724/SP.J.1145.2010.00730]
 YANG Junshi,ZHOU Houzhen,LI Guoxin,et al.Treatment of Wastewater from Alcohol Fuel Production with Sweet Potato by Combined Process[J].Chinese Journal of Applied & Environmental Biology,2010,16(02):730.[doi:10.3724/SP.J.1145.2010.00730]
[4]李岩,王海燕,张义正.甘薯蔗糖转运蛋白IbSUT1x在酵母细胞中的定位[J].应用与环境生物学报,2010,16(06):798.[doi:10.3724/SP.J.1145.2010.00798]
 LI Yan,WANG Haiyan,ZHANG Yizheng.Localization of IbSUT1x Protein from Ipomoea batatas (L.) Lam in Yeast Cells[J].Chinese Journal of Applied & Environmental Biology,2010,16(02):798.[doi:10.3724/SP.J.1145.2010.00798]
[5]段鹏,张义正,谭雪梅.甘薯块根腐烂真菌的分离和鉴定[J].应用与环境生物学报,2011,17(02):260.[doi:10.3724/SP.J.1145.2011.00260]
 DUAN Peng,ZHANG Yizheng,TAN Xuemei.Isolation and Identification of Fungi from Rotted Tuber of Sweet Potato[J].Chinese Journal of Applied & Environmental Biology,2011,17(02):260.[doi:10.3724/SP.J.1145.2011.00260]
[6]王广珺,何明雄,张义正.运动发酵单胞菌共表达α-淀粉酶和葡萄糖淀粉酶发酵甘薯生产乙醇[J].应用与环境生物学报,2012,18(05):785.[doi:10.3724/SP.J.1145.2012.00785]
 WANG Guangjun,HE Mingxiong,ZHANG Yizheng.Co-expression of α-amylase and Glucoamylase in Zymomonas mobilis and Direct Ethanol Production from Sweet Potato[J].Chinese Journal of Applied & Environmental Biology,2012,18(02):785.[doi:10.3724/SP.J.1145.2012.00785]
[7]陈娇,姜玉松,张义正,等.甘薯LEA2基因的克隆与表达分析[J].应用与环境生物学报,2014,20(02):204.[doi:10.3724/SP.J.1145.2014.00204]
 CHEN Jiao,JIANG Yusong,ZHANG Yizheng,et al.Cloning and expression analyses of LEA2 gene from Ipomoea batatas[J].Chinese Journal of Applied & Environmental Biology,2014,20(02):204.[doi:10.3724/SP.J.1145.2014.00204]

备注/Memo

备注/Memo:
国家科技支撑计划项目(2007BAD78B03)和四川省“十一五”重点科技攻关项目(07SG111-003-1)资助
更新日期/Last Update: 2013-05-02