期刊文献+

斑马鱼心脏标记基因VMHC多抗血清的制备 被引量:2

Preparation of Polyclonal Antiserum of Heart Marker Gene VMHC in Zebrafish
下载PDF
导出
摘要 斑马鱼基因VMHC(Ventricular myosin heavy chain)是心脏发育早期的标志基因,制备该基因多抗血清,首先通过生物信息学方法,选择VMHC基因中特异性强、具亲水性的一段核苷酸序列(3 772~4 221 bp),通过PCR扩增,将片段重组到原核表达载体pGEX-4T-1,通过原核诱导表达获得含有表达目的片段的融合蛋白,以该融合蛋白免疫小鼠,获得VMHC多克隆抗鼠血清.对该多抗血清抗体进行验证,结果具有很好特异性和较高效价,可以用作western-blot、免疫印迹等试验分析. Zebrafish gene VMHC (Ventricular myosin heavy chain) is a marker gen in the early development heart. In order to get the polyclonal antiserum of VMHC, a VMHC-specifie, hydrophilie nucleotide sequence ( CDS from 3 772 to 4 221 bp) was selected by using bioinformatics methods, and the sequence was amplified by PCR. The PCR product was inserted into the prokaryotic expression vector pGEX-4T-1 and the fusion protein was induced to express in BL21 cells. The mouse was immunized by injecting appropriate amount of fusion protein and the VMHC polyclonal antiserum was obtained. The antiserum's titer and specification was identified. The results showed that the VMHC polyclonal antiserum was practical and available for western-blotting analysis and immueprecipitation.
出处 《湖南师范大学自然科学学报》 CAS 北大核心 2011年第2期71-74,共4页 Journal of Natural Science of Hunan Normal University
基金 湖南省普通高校学科带头人培养计划资助项目(湘教通2008-315) 国家精品课程资助项目(教高函2009-21)
关键词 VMHC 标记基因 融合蛋白 多抗血清 VMHC marker gen fusion protein polyelonal antiserum
  • 相关文献

参考文献9

  • 1WEISS A, LEINWAND L A. The mammalian myosin heavy chain gene family [ J ]. Annu Rev Cell Dev Biol, 1996,12:417-439.
  • 2MORKIN E. Control of cardiac myosin heavy chain gene expression[J]. Microsc Res Tech,2000 ,50 :522-531.
  • 3YELON D, HORNE S A, STAINIER D Y. Restricted expression of cardiac myosin genes reveals regulated aspects of heart tube assembly in zebrafish[ J]. Dev Biol, 1999,21d :23-37.
  • 4BERDOUGO E, COLEMAN H, LEE D H, et al. Mutation of weak Atrium/atrial myosin heavy chain disrupts atrial function and influences ventricnlar morphogenesis in zebrafish[ J ]. Development,2003,130:6121-6129.
  • 5BRANT M W, DEREK L S, WOLFGANG D, et al. Gridlock, a localized heritable vascular patterning defect in the zebrafish [J]. Nat Med, 1995,1:1143-1147.
  • 6Sam Brook J,Russel D W.分子克隆实验指南[M].黄培堂,译.3版.北京:科学出版社,2002,463-471.
  • 7YUAN C, REULAND J M, LEE L, et al. Optimized expression and refolding of human kerato epithelin in BI21 (DE3) [J]. Protein Expr Purif, 2004,35:39-45.
  • 8ODENTHAL J, ROSSNAGEL K, HAFFTER P, et al. Mutations affecting xanthophore pigmentation in the zebrafish, Danio re- rio [ J ]. Development, 1996,123 : 391-398.
  • 9王华,张云,吴秀山,袁婺洲.斑马鱼心脏标记基因pitx2的多克隆抗体的制备[J].湖南师范大学自然科学学报,2010,33(2):98-101. 被引量:2

二级参考文献9

  • 1BAKKERS J,VERHOEVEN M C,SEYFRIED S A.Shaping the zebrafish heart:from left-fight axis specil cation to epithelial tissue morphogenesis[J].Developmental Biology,2009,330:213-220.
  • 2HARVEY R P.Cardiac looping-an uneasy deal with laterality[J].Developmental Biology,1998,9(1):101-108.
  • 3NONAKA S,SHIRATORI H,SAIJOH Y,et al.Determination of left-right patterning of the mouse embryo by artificial nodal flow[J].Nature,2002,418:96-99.
  • 4JEFFREY J,ESSNER W W,BRANFORD J Z,et al.Mesendoderm and left-right brain,heart and gut development are differentially regulated by pitx2 isoforms[J].Development,2000,127:1081-1093.
  • 5MARINA C,HERBERT S,AXEL S,et al.The homeobox gene pitx2:mediator of asymm-etric left-right signaling in vertebrate heart and gut looping[J].Development,1999,126:1225-1234.
  • 6CAMPIONE M,ROS M A,ICARDO J M,et al.Pitx2 expression defines a left cardiac lineage of cells:evidence for atrial and ventricular molecular isomerism in the iv/iv mice[J].Developmental Biology,2001,231 (1):252-264.
  • 7RYAN A K,BLUNBERG B,RODRIGUEZ E C,et al.Pitx2 determines loft-right asymmetry of internal organs in vertebrates[J].Nature,1998,394(6693):545-551.
  • 8DI A,WEI L,LI J M,et al.Pitx2 regulates cardiac left-right asymmetry by patterning second cardiac lineage-derived myocardium[J].Developmental Biology,2006,296:437-449.
  • 9刘冀珑,谭燕.左右不对称信号分子Pitx2[J].生物化学与生物物理进展,1999,26(1):3-5. 被引量:2

共引文献67

同被引文献15

  • 1VALLEJO M,RON D,MILLER C P.C/ATF,a member of the activating transcription factor family of DNA-binding proteins, dimerizes with CAAT/enhancer-binding proteins and directs their binding to cAMP response elements[J].Proc Nail Acad Sci USA,1993,90(10):4679-4683.
  • 2LIAN G,HAI T.Characterization of human activating transcription factor 4,a transcriptional activator that interacts with multiple domains of cAMP-responsive element-binding protein (CREB)-binding protein[J].J Biol Chem,1997,272(38): 24088-24095.
  • 3BAGHERI-YARMAND R,VADLAMUDI R K,KUMAR R.Activating transcription factor 4 over expression inhibits proliferation and differentiation of mammary epithelium resulting in impaired lactation and accelerated involution[J].J Biol Chem,2003, 278(19):17421-17429.
  • 4ZHANG X,YU S,GALSON D L,et al.Activating transcription factor 4 is critical for proliferation and survival in primary bone marrow stromal cells and calvarial osteoblasts[J].J Cell Biochem, 2008,105(3):885-895.
  • 5YUAN C,REULAND J M,LEE L,et al.Optimized expression and refolding of human kerato epithelin in BL21(DE3 )[J]. Protein Expression and Purification,2004,35:39-45.
  • 6ODENTHAL J,ROSSNAGEL K,HAFFTER P,et al.Mutations affecting xanthoph-ore pigmentation in the Zebrafish,Danio rerio [J].Development,1996,123:391-398.
  • 7ZHANG Ning, LIU Jing, DING Xia, et al. Fbxl5 interacts with p150 Glued and regulates its ubiquitination [ J]. Biochemical and Biophysical Research Communications, 2007, 359:34-39.
  • 8GAGNE J M, DOWNES B P, SHIU S H, et al. The F-box es- ubunit of the SCF E3 complex is encoded by a diverse superfami- lyof genes in Arabidopsis [ J]. Proc Natl Acad Sci, USA 2002, 99 ( 17 ) :11519-11524.
  • 9KAREN L CRAIG, MIKE TYERS. The F-box: a new motif for ubiquitin dependent proteolysis in cell cycle regulation and signaltransduction[J]. Progress in Biophysics & Molecular Biology, 1999, 72:299-328.
  • 10SCHRAMMEUER B, RISSEEUW E, PANSEGRAU W, et al. Interaction of the virulence protein VirF of Agrobaeterium tumefa- ciens with plant homologs of the yeast Skpl protein[ J]. Curt Bi- ol, 2001, 11(4) :258-62.

引证文献2

二级引证文献2

相关作者

内容加载中请稍等...

相关机构

内容加载中请稍等...

相关主题

内容加载中请稍等...

浏览历史

内容加载中请稍等...
;
使用帮助 返回顶部