酵母双杂交技术鉴定与拟南芥AtMT-A70B相互作用基因探讨

论文片段—ceofMB*/pENTR27-30Chapter4ScreeningArobidopsiscDNApbrarybyMB*bait30-414.1TranerMB*topDEST32usinggatewaytechnology304.2TestoftheautoactivationoftheMB*-bait30-314.3Library-scaletranormationofyeastcompetentcellM20331-324.4Identificationofpositiveclones32-414.4.拟南芥论文,RT-PCR论文,基因文库论文,酵母双杂交论文,转移酶论文,
摘要：mRNA在线粒体内翻译成蛋白质论文网站大全。前体mRNA在线粒体之前经过一系列转录后修饰,这些修饰包括在5′端加上帽子结构,3′端加上多聚腺苷酸尾巴以及剪接修饰。除了这些常见的修饰外,对特定核苷酸的修饰也是经常发生的怎么写论文。RNA分子生物学研究RNA的转录后修饰在调控基因表达发挥着的作用。尤其是腺嘌呤上第六个氮原子的化在mRNA的加工和运输中的研究越来越该研究的热点问题。负责拟南芥中腺嘌呤化酶(AtMT-A70)的基因(MB~*)已经被Rupert.G.Fray博士的实验小组分离出来,的研究该基因对拟南芥种子胚胎发育致命的影响。为了寻找完成化所的其他成员,本试验研究用RT-PCR和PCR技术对MB~*了克隆,并用其作为探针在酵母双杂交系统下来扫描拟南芥花药cDNA基因文库。经过反复筛选证明,克隆B1/2,B1/14,B1/20,B13/1和B15/7与MB~*有强烈的作用。对这些克隆测序和Blast分析证明克隆B15/7,B1/14,B1/20表达的蛋白在拟南芥中的代号为At5g49020,但是功能未知;克隆B13/1,B1/2表达的蛋白在拟南芥中的代号是At3g61690,其功能为精氨酸-N-转移酶;它们都参与化,该发现尚属首次。本试验结果对认识mRNA化以及参与这一的蛋白具有的硕士毕业论文。在试验中用到了高级的生物学技术,比如说PCR,RT-PCR和酵母双杂交技术。关键词：拟南芥论文RT-PCR论文基因文库论文酵母双杂交论文转移酶论文
Abstract5-6
Abbreviations6-7
Chapter 1 Introduction7-13
1.1 About the most common modification of adenosines in mRNA7-8

1.2 About the MB*(AtMT-A70B)8

1.3 About the gateway cloning technology8-9

1.4 About the Yeast two Hybrid system9-10

1.5 The ProQuest? Two-Hybrid System with Gateway? Technology10-12

1.6 The procedure of the experiment12-13

Chapter 2 Material and methods13-27

2.1 Growth of plant13

2.2 RNA extraction13-14

2.3 Preparation of agarose gels14

2.4 RT-PCR(Reverse Transcription PCR)14-15
2.5 QIaquick gel extraction kit protocol15
2.6 Information of pENTR/D-TOPO vector15-16
2.7 Put PCR products(MB*) into Entry vector use Gateway? Cloning Technology(BP Recombination reaction)16-17
2.8 Preparation of competent host strain DH5a E. cop cells17-18
2.9 Tranormation of competent DH5a E. cop cells by MB* / pENTR18
2.10 MB*/ pENTR plaid DNA minipreps(QIAGEN)18
2.11 Mediums for Yeast-two Hybrid screen18-20

2.12 Plates for Yeast two Hybrid screen20

2.13 Low-Copy-Number DB and AD Vectors (ARS/CEN)20-22

2.14 Small-Scale Tranormation of MaV20322

2.15 Library scale tranormation22-23

2.16 Galactosidase filter assay23

2.17 Assessing Self-Activation23-24

2.18 Traner MB* to pDESTTM 32 and two-hybrid screen24-25

2.19 Yeast plaid isolation25-27

Chapter 3 Cloning of MB*27-30

3.1 Cloning of MB*27

3.2 Construction of the bait plaid27

3.3 Sequence of MB*/ pENTR27-30

Chapter 4 Screening Arobidopsis cDNA pbrary by MB* bait30-41
4.1 Traner MB* to pDEST32 using gateway technology30
4.2 Test of the autoactivation of the MB*-bait30-31
4.3 Library-scale tranormation of yeast competent cell M20331-32
4.4 Identification of positive clones32-41

4.1 Growth assays32

4.2 X-gal assays32-33

4.3 Library plaid isolation33-34

4.4 Confirmation of positive clones34-35

4.4.5 Sequence of the pos论文片段—itiveclones35-404.4.6Identificationofpositiveclones(BLASTsearches)40-41Chapter5Discussionandfuturework41-43Reference43-46摘要46-47上一页12拟南芥论文,RT-PCR论文,基因文库论文,酵母双杂交论文,转移酶论文,
itive clones35-40
4.4.6 Identification of positive clones (BLAST searches)40-41
Chapter 5 Discussion and future work41-43
Reference43-46
摘要46-47