Scientia Agricultura Sinica ›› 2012, Vol. 45 ›› Issue (20): 4279-4287.doi: 10.3864/j.issn.0578-1752.2012.20.018

• ANIMAL SCIENCE·RESOURCE INSECT • Previous Articles     Next Articles

Cloning and Activity Analysis of a Midgut-Specific Promoter BmAPN in Silkworm (Bombyx mori)

 LU  Gai, CHENG  Ting-Cai, JIANG  Liang, JIN  Sheng-Kai, LIN  Ping, HU  Cui-Mei, XIA  Qing-You   

  1. 1.西南大学蚕学与系统生物学研究所/家蚕基因组生物学国家重点实验室，重庆 400716
  • Received:2012-02-13 Online:2012-10-15 Published:2012-03-23

PDF

894

Cited

2

Abstract: 【Objective】The objective of this study is to provide a new tool for the immune-response and application research, and to clone and identify a midgut-specific promoter from Bombyx mori.【Method】 The 5′ upstream regulatory sequence of midgut-specific gene BmAPN (GenBank accession: BAA33715.1) was obtained by PCR from B. mori genomic DNA. A transgenic expression vector named pBac[BmAPN-EGFP-SV40, 3×P3-DsRed] was constructed, in which the reporter gene EGFP was driven by the BmAPN promoter. Furthermore, transgenic B. mori lines was obtained by microinjection and the activity of the promoter at the individual level were detected.【Result】The EGFP driven by a 1 598 bp BmAPN promoter was just specifically expressed in B. mori midgut. It was consistent with the expression pattern of endogenous gene BmAPN. The promoter activity was higher in B. mori larvae than other stages. And the activity in newly exuviated larvae was higher than that of molting larvae. It was speculated that specific components of the promoter might be regulated by hormone.【Conclusion】The BmAPN promoter is an active and midgut-specific promoter in B. mori.

Key words: Bombyx mori, midgut-specific, promoter, transgene

[1]Hua G, Tsukamoto K, Rasilo M L, Ikezawa H. Molecular cloning of a GPI-anchored aminopeptidase N from Bombyx mori midgut : a putative receptor for Bacillus thuringiensis CryIA toxin. Gene, 1998, 214 (1/2): 177-185.

[2]Yaoi K, Nakanishi K, Kadotani T, Imamura M, Koizumi N, Iwahana H, Sato R. cDNA cloning and expression of Bacillus thuringiensis Cry1Aa toxin binding 120 kDa aminopeptidase N from Bombyx mori. Biochimica et Biophysica Acta, 1999, 1444 (1): 131-137.

[3]Crava C M, Bel Y, Lee S F, Manachini B, Heckel D G, Escriche B. Study of the aminopeptidase N gene family in the lepidopterans Ostrinia nubilalis (Hübner) and Bombyx mori (L.): sequences, mapping and expression. Insect Biochemistry and Molecular Biology, 2010, 40(7): 506-515.

[4]Vadlamudi R K, Weber E, Ji I, Ji T H, Bulla Jr. L A. Cloning and expression of a receptor for an insecticidal toxin of Bacillus thuringiensis. Journal of Biological Chemistry, 1995, 270(10): 5490-5494.

[5]Pacheco S, Gómez I, Arenas I, Saab-Rincon G, Rodríguez-Almazán C, Gill S S, Bravo A, Soberón M. Domain II loop 3 of Bacillus thuringiensis Cry1Ab toxin is involved in a "ping pong" binding mechanism with Manduca sexta aminopeptidase-N and cadherin receptors. The Journal of Biological Chemistry, 2009, 284(47): 32750-32757.

[6]Bravo A, Gómez I, Conde J, Muñoz-Garay C, Sánchez J, Miranda R, Zhuang M, Gill S S, Soberón M. Oligomerization triggers binding of a Bacillus thuringiensis Cry1Ab pore-forming toxin to aminopeptidase N receptor leading to insertion into membrane microdomains. Biochimica et Biophysica Acta, 2004, 1667(1): 38-46.

[7]Bravo A, Gill S S, Soberón M. Mode of action of Bacillus thuringiensis Cry and Cyt toxins and their potential for insect control. Toxicon, 2007, 49(4): 423-435.

[8]Bravo A, Likitvivatanavong S, Gill S S, Soberón M. Bacillus thuringiensis: A story of a successful bioinsecticide. Insect Biochemistry and Molecular Biology, 2011, 41(7): 423-431.

[9]Rajagopal R, Sivakumar S, Agrawal N, Malhotra P, Bhatnagar R K. Silencing of midgut aminopeptidase N of Spodoptera litura by double-stranded RNA establishes its role as Bacillus thuringiensis toxin receptor. The Journal of Biological Chemistry, 2002, 277(49): 46849-46851.

[10]Herrero S, Gechev T, Bakker P L, Moar W J, de Maagd R A. Bacillus thuringiensis Cry1Ca-resistant Spodoptera exigua lacks expression of one of four aminopeptidase N genes. BMC Genomics, 2005, 6: 96.

[11]Abraham E G, Donnelly-Doman M, Fujioka H, Ghosh A, Moreira L, Jacobs-Lorena M. Driving midgut-specific expression and secretion of a foreign protein in transgenic mosquitoes with AgAper1 regulatory elements. Insect Molecular Biology, 2005, 14(3): 271-279.

[12]Zhao A C, Zhao T F, Zhang Y S, Xia Q Y, Lu C, Zhou Z Y, Xiang Z H, Nakagaki M. New and highly efficient expression systems for expressing selectively foreign protein in the silk glands of transgenic silkworm. Transgenic Research, 2010, 19(1): 29-44.

[13]邓党军, 徐汉福, 王  峰, 马三垣, 夏庆友. BmLSP基因启动子驱动DsRed在转基因家蚕中的表达分析. 蚕业科学, 2011, 37(2): 200-205.

Deng D J, Xu H F, Wang F, Ma S Y, Xia Q Y. Expression analysis of DsRed gene driven by promoter of BmLSP gene in transgenic silkworm, Bombyx mori. Science of Sericulture, 2011, 37(2): 200-205. (in Chinese)

[14]胡翠美, 王  菲, 宋  亮, 夏庆友. 家蚕中肠cDNA T7噬菌体展示文库的构建和免疫相关基因的淘选. 蚕业科学, 2011, 37(4): 642-649.

Hu C M, Wang F, Song L, Xia Q Y. Construction of a T7 phage display cDNA library and panning for immune-related genes from midgut of silkworm, Bombyx mori. Science of Sericulture, 2011, 37(4): 642-649. (in Chinese)

[15]马三垣, 徐汉福, 段建平, 赵爱春, 张美蓉, 夏庆友. 家蚕转基因技术中若干因素对转基因效率的影响. 昆虫学报, 2009, 52(6): 595-603.

Ma S Y, Xu H F, Duan J P, Zhao A C, Zhang M R, Xia Q Y. Effect of several factors on the transformation efficiency in the transgenic technology of silkworm, Bombyx mori. Acta Entomologica Sinica, 2009, 52(6): 595-603. (in Chinese)

[16]Wang G H, Xia Q Y, Cheng D J, Duan J, Zhao P, Chen J, Zhu L. Reference genes identified in the silkworm Bombyx mori during metamorphism based on oligonucleotide microarray and confirmed by qRT-PCR. Insect Science, 2008, 15(5): 405-413.

[17]Livak K J, Schmittgen T D. Analysis of relative gene expression data using real-time quantitative PCR and the 2－ΔΔCt method. Methods, 2001, 25(4): 402-408.

[18]Fernandes M, Xiao H, Lis J T. Fine structure analyses of the Drosophila and Saccharomyces heat shock factor-heat shock element interactions. Nucleic Acids Research, 1994, 22(2): 167-173.

[19]Bray S J, Kafatos F C. Developmental function of Elf-1: an essential transcription factor during embryogenesis in Drosophila. Genes & Development, 1991, 5(9): 1672-1683.

[20]Akiyama Y, Hosoya T, Poole A M, Hotta Y. The gcm-motif: a novel DNA-binding motif conserved in Drosophila and mammals. Proceedings of the National Academy of Sciences of the United States of America, 1996, 93(25): 14912-14916.

[21]Lecourtois M, Schweisguth F. The neurogenic suppressor of hairless DNA-binding protein mediates the transcriptional activation of the enhancer of split complex genes triggered by notch signaling. Genes & Development, 1995, 9(21): 2598-2608.

[22]Bailey A M, Posakony J W. Suppressor of hairless directly activates transcription of enhancer of split complex genes in response to notch receptor activity. Genes & Development, 1995, 9(21): 2609-2622.

[23]Ekker S C, von Kessler D P, Beachy P A. Differential DNA sequence recognition is a determinant of specificity in homeotic gene action. The EMBO Journal, 1992, 11(11): 4059-4072.

[24]Von Kalm L, Crossgrove K, Von Seggern D, Guild G M, Beckendorf S K. The broad-complex directly controls a tissue-specific response to the steroid hormone ecdysone at the onset of Drosophila metamorphosis. The EMBO Journal, 1994, 13(15): 3505-3516.

[25]Stanojevi? D, Hoey T, Levine M. Sequence-specific DNA-binding activities of the gap proteins encoded by hunchback and Krueppel in Drosophila. Nature, 1989, 341(6240): 331-335.

[26]Treisman J, Desplan C. The products of the Drosophila gap genes hunchback and Krueppel bind to the hunchback promoters. Nature, 1989, 341(6240): 335-337.

[27]Read D, Manley J L. Alternatively spliced transcripts of the Drosophila tramtrack gene encode zinc finger proteins with distinct DNA binding specificities. The EMBO Journal, 1992, 11(3): 1035-1044.

[28]Brinster R L, Allen J M, Behringer R R, Gelinas R E, Palmiter R D. Introns increase transcriptional efficiency in transgenic mice. Proceedings of the National Academy of Sciences of the United States of America, 1988, 85(3): 836-840.

[29]Rafiq M, Suen C K, Choudhury N, Joannou C L, White K N, Evans R W. Expression of recombinant human ceruloplasmin-an absolute requirement for splicing signals in the expression cassette. FEBS Letters, 1997, 407(2): 132-136.

[30]Wang H B, Iwanaga M, Kawasaki H. Activation of BMWCP10 promoter and regulation by BR-C Z2 in wing disc of Bombyx mori. Insect Biochemistry and Molecular Biology, 2009, 39(9): 615-623.

[31]Wang H B, Nita M, Iwanaga M, Kawasaki H. BetaFTZ-F1 and broad-complex positively regulate the transcription of the wing cuticle protein gene, BMWCP5, in wing discs of Bombyx mori. Insect Biochemistry and Molecular Biology, 2009, 39(9): 624-633.

[32]Yaoi K, Nakanishi K, Kadotani T, Imamura M, Koizumi N, Iwahana H, Sato R. Bacillus thuringiensis Cry1Aa toxin-binding region of Bombyx mori aminopeptidase N. FEBS Letters, 1999, 463(3): 221-224.

[33]Nakanishi K, Yaoi K, Nagino Y, Hara H, Kitami M, Atsumi S, Miura N, Sato R. Aminopeptidase N isoforms from the midgut of Bombyx mori and Plutella xylostella - their classification and the factors that determine their binding specificity to Bacillus thuringiensis Cry1A toxin. FEBS Letters, 2002, 519(1/3): 215-220.
[1] LI XuFei,YANG ShengDi,LI SongQi,LIU HaiNan,PEI MaoSong,WEI TongLu,GUO DaLong,YU YiHe. Analysis of VlCKX4 Expression Characteristics and Prediction of Transcriptional Regulation in Grape [J]. Scientia Agricultura Sinica, 2023, 56(1): 144-155.
[2] LONG YanBi,WU YunFei,ZHANG Qian,CHEN Peng,PAN MinHui. Screening and Identification of HSP90 Interacting Proteins in Silkworm (Bombyx mori) [J]. Scientia Agricultura Sinica, 2022, 55(6): 1253-1262.
[3] YAN Qiang,XUE Dong,HU YaQun,ZHOU YanYan,WEI YaWen,YUAN XingXing,CHEN Xin. Identification of the Root-Specific Soybean GmPR1-9 Promoter and Application in Phytophthora Root-Rot Resistance [J]. Scientia Agricultura Sinica, 2022, 55(20): 3885-3896.
[4] XU XianBin,GENG XiaoYue,LI Hui,SUN LiJuan,ZHENG Huan,TAO JianMin. Transcriptome Analysis of Genes Involved in ABA-Induced Anthocyanin Accumulation in Grape [J]. Scientia Agricultura Sinica, 2022, 55(1): 134-151.
[5] LIU Chuang,GAO Zhen,YAO YuXin,DU YuanPeng. Functional Identification of Grape Potassium Ion Transporter VviHKT1;7 Under Salt Stress [J]. Scientia Agricultura Sinica, 2021, 54(9): 1952-1963.
[6] SHA RenHe,LAN LiMing,WANG SanHong,LUO ChangGuo. The Resistance Mechanism of Apple Transcription Factor MdWRKY40b to Powdery Mildew [J]. Scientia Agricultura Sinica, 2021, 54(24): 5220-5229.
[7] DU Xing,ZENG Qiang,LIU Lu,LI QiQi,YANG Liu,PAN ZengXiang,LI QiFa. Identification of the Core Promoter of Linc-NORFA and Its Transcriptional Regulation in Erhualian Pig [J]. Scientia Agricultura Sinica, 2021, 54(15): 3331-3342.
[8] WANG Xiao,CAI Jian,ZHOU Qin,DAI TingBo,JIANG Dong. Physiological Mechanisms of Abiotic Stress Priming Induced the Crops Stress Tolerance: A Review [J]. Scientia Agricultura Sinica, 2021, 54(11): 2287-2301.
[9] CHEN Liu,NI Zheng,YU Bin,HUA JiongGang,YE WeiCheng,YUN Tao,LIU KeShu,ZHU YinChu,ZHANG Cun. Optimized Promoter Regulating of Duck Tembusu Virus E Protein Expression Delivered by a Vectored Duck Enteritis Virus in vitro [J]. Scientia Agricultura Sinica, 2020, 53(24): 5125-5134.
[10] CHEN Peng,BAO XiYan,KANG TaoTao,DONG ZhanQi,ZHU Yan,PAN MinHui,LU Cheng. Screening and Identification of Proteins Interacting with Bombyx mori IAP and Their Effects on BmNPV Proliferation [J]. Scientia Agricultura Sinica, 2019, 52(3): 558-567.
[11] BIAN ShuXun,HAN XiaoLei,YUAN GaoPeng,ZHANG LiYi,TIAN Yi,ZHANG CaiXia,CONG PeiHua. Cloning and Functional Analysis of U6 Promoter in Apple [J]. Scientia Agricultura Sinica, 2019, 52(23): 4364-4373.
[12] DONG ZhanQi,JIANG YaMing,PAN MinHui. Screening and Identification of Candidate Proteins Interacting with BmHSP60 in the Silkworm (Bombyx mori) [J]. Scientia Agricultura Sinica, 2019, 52(2): 376-384.
[13] GE Ting,HUANG Xue,XIE RangJin. Cloning, Subcellular Localization and Expression Analysis of CitPG34 in Citrus [J]. Scientia Agricultura Sinica, 2019, 52(19): 3404-3416.
[14] YI Min,LÜ Qing,LIU KeKe,WANG LiJun,WU YuJiao,ZHOU ZeYang,LONG MengXian. Expression, Purification and Localization Analysis of Polar Tube Protein 2 (NbPTP2) from Nosema bombycis [J]. Scientia Agricultura Sinica, 2019, 52(10): 1830-1838.
[15] ZHANG Kui,LI ChongYang,SU JingJing,TAN Juan,XU Man,CUI HongJuan. Expression, Purification and Immunologic Function of Integrin β2 in the Silkworm (Bombyx mori) [J]. Scientia Agricultura Sinica, 2019, 52(1): 181-190.
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