pBD-GAL4-Cam

编号	载体名称
YB2870	pBD-GAL4-Cam

 

pBD-GAL4-Cam载体基本信息

载体名称:	pBD-GAL4-Cam
质粒类型:	酵母双杂交载体；酵母噬粒载体
克隆方法:	多克隆位点，限制性内切酶
启动子:	ADH1
载体大小:	6491 bp
5' 测序引物及序列:	--
3' 测序引物及序列:	--
载体标签:	无
载体抗性:	氯霉素Chloramphenicol
真核筛选标记:	TRP1
克隆菌株:	XL1-Blue MRF'
表达菌株:	YRG-2
备注:	含a beta-gal报告基因。配套载体为pAD-GAL4-2.1
产品目录号:	--
组成型/诱导型:	组成型

pBD-GAL4-Cam载体质粒图谱和多克隆位点信息

pBD-GAL4-Cam载体简介

酵母双杂交系统Protein–protein interactions occur in many biological processes including replication, transcription, secretion, signal transduction, and metabolism. A fundamental question in the study of any protein is to identify proteins that interact with a given protein in vivo. Intense research efforts are focused on the identification of these proteins. The GAL4 two-hybrid phagemid vector system, a eukaryotic system to detect protein–protein interactions in vivo, provides a method for the rapid identification of genes encoding proteins that interact with a given protein (i.e., a bait protein).1,2 The system is based on the ability to separate eukaryotic transcriptional activators into two separate domains, the DNAbinding domain (BD) and the transcriptional activation domain (AD). In theGAL4 two-hybrid phagemid vector system, proteins that interact with the bait protein are identified by generating hybrids of the yeast GAL4 BD and the bait protein (X) and the GAL4 AD and a library of proteins (Y). Neither hybrid protein is capable of initiating specific transcription of reporter genes in yeast in the absence of a specific interaction with the other hybrid protein. When the hybrid protein X is expressed in yeast, the GAL4 BD binds X to specific DNA sequences in the yeast chromosome defined by the GAL1 or GAL4 upstream activating sequences (UASGAL1 or UASGAL4, respectively), which regulate the expression of a reporter gene. Binding of X to the UAS is not sufficient to initiate transcription of the reporter gene. When Y is expressed in yeast, the AD interacts with other components of the transcription machinery required to initiate transcription of the reporter gene. However, Y alone is not localized to the reporter gene UAS and therefore does not activate transcription of the reporter gene. When a specific interaction between X and Y localizes both the GAL4 BD and GAL4 AD to the reporter gene UAS, transcriptional activation of the reporter gene occurs (Figure 2B). The reporter genes in the GAL4 twohybrid phagemid vector system are β-galactosidase (lacZ) and histidine (HIS3). 双杂交载体pAD-GAL4-2.1和pBD-GAL4-CamThe pAD-GAL4-2.1 phagemid vector contains a multiple cloning site (MCS) with BamH I, Nhe I, EcoR I, Xho I, Sal I, Xba I, Pst I, and Bgl II restriction sites. The pBD-GAL4-Cam phagemid vector contains an MCS with EcoR I, Srf I, Sma I, Xho I, Sal I, Xba I, and Pst I restriction sites. The unique EcoR I and Xho I cloning sites in the pAD-GAL4-2.1 vector make this vector compatible with the Agilent cDNA Synthesis Kit for the preparation of unidirectional cDNA libraries. The unique EcoR I and Sal I cloning sites are used for the preparation of cDNA libraries in the pBD-GAL4 Cam phagemid vector because the Xho I site in the MCS is not unique. The unique BamH I, Nhe I, and EcoR I sites at the 5′ end and the Xho I, Sal I, Xba I, and Bgl II sites at the 3′ end of the DNA insert facilitate the transfer of DNA encoding the target protein into commonly used protein expression/purification vectors. Genomic DNA digested with Mbo I, BamH I, or Sau3A I and partially filled-in can be inserted into a partially filled-in Xho I site in the pAD-GAL4-2.1 phagemid vector. The Xba I site in the pAD-GAL4-2.1 and pBD-GAL4-Cam phagemid vectors is not unique and contains the UAG amber suppressor in the same translational reading frame as the GAL4 domain. DNA should therefore be inserted such that the Xba I site is not between the GAL4 domain and the DNA insert. The pAD-GAL4-2.1 and pBD-GAL4 Cam phagemid vectors contain the pUC origin for replication and an f1 origin for production of single-stranded DNA (ssDNA) in E. coli. Single-stranded DNA can be used for DNA sequencing or site-directed mutagenesis. The pAD-GAL4-2.1 and pBDGAL4-Cam phagemid vectors contain ampicillin-resistance gene [β-lactamase (bla)] and chloramphenicol acetyltransferase genes, respectively, for selection with ampicillin and chloramphenicol in E. coli. The pADGAL4-2.1 and pBD-GAL4 Cam phagemid vectors contain the 2μ origin for replication. For selection in yeast, the pAD-GAL4-2.1 phagemid vector contains the LEU2 gene and the pBD-GAL4 Cam phagemid vector contains the TRP1 gene. The hybrid protein is expressed by the alcohol dehydrogenase 1 (ADH1) promoter (P ADH1) and is terminated by the ADH1 terminator (T ADH1). 

pBD-GAL4-Cam载体序列

pBD-GAL4-Cam, 6491 bp                          version 045002 NOTE: The following sequence has been verified for accuracy at the junctions. The remainder of the sequence has been obtained from existing data.      1  GCTTGCATGC AACTTCTTTT CTTTTTTTTT CTTTTCTCTC TCCCCCGTTG    51  TTGTCTCACC ATATCCGCAA TGACAAAAAA ATGATGGAAG ACACTAAAGG   101  AAAAAATTAA CGACAAAGAC AGCACCAACA GATGTCGTTG TTCCAGAGCT   151  GATGAGGGGT ATCTCGAAGC ACACGAAACT TTTTCCTTCC TTCATTCACG   201  CACACTACTC TCTAATGAGC AACGGTATAC GGCCTTCCTT CCAGTTACTT   251  GAATTTGAAA TAAAAAAAAG TTTGCTGTCT TGCTATCAAG TATAAATAGA   301  CCTGCAATTA TTAATCTTTT GTTTCCTCGT CATTGTTCTC GTTCCCTTTC   351  TTCCTTGTTT CTTTTTCTGC ACAATATTTC AAGCTATACC AAGCATACAA   401  TCAACTCCAA GCTTGAAGCA AGCCTCCTGA AAGATGAAGC TACTGTCTTC   451  TATCGAACAA GCATGCGATA TTTGCCGACT TAAAAAGCTC AAGTGCTCCA   501  AAGAAAAACC GAAGTGCGCC AAGTGTCTGA AGAACAACTG GGAGTGTCGC   551  TACTCTCCCA AAACCAAAAG GTCTCCGCTG ACTAGGGCAC ATCTGACAGA   601  AGTGGAATCA AGGCTAGAAA GACTGGAACA GCTATTTCTA CTGATTTTTC   651  CTCGAGAAGA CCTTGACATG ATTTTGAAAA TGGATTCTTT ACAGGATATA   701  AAAGCATTGT TAACAGGATT ATTTGTACAA GATAATGTGA ATAAAGATGC   751  CGTCACAGAT AGATTGGCTT CAGTGGAGAC TGATATGCCT CTAACATTGA   801  GACAGCATAG AATAAGTGCG ACATCATCAT CGGAAGAGAG TAGTAACAAA   851  GGTCAAAGAC AGTTGACTGT ATCGCCGGAA TTCGCCCGGG CCTCGAGCCC   901  GGGTCGACTC TAGAGCCCTA TAGTGAGTCG TATTACTGCA GCCAAGCTAA   951  TTCCGGGCGA ATTTCTTATG ATTTATGATT TTTATTATTA AATAAGTTAT  1001  AAAAAAAATA AGTGTATACA AATTTTAAAG TGACTCTTAG GTTTTAAAAC  1051  GAAAATTCTT GTTCTTGAGT AACTCTTTCC TGTAGGTCAG GTTGCTTTCT  1101  CAGGTATAGC ATGAGGTCGC TCTTATTGAC CACACCTCTA CCGGCATGCC  1151  GGCAAGTGCA CAAACAATAC TTAAATAAAT ACTACTCAGT AATAACCTAT  1201  TTCTTAGCAT TTTTGACGAA ATTTGCTATT TTGTTAGAGT CTTTTACACC  1251  ATTTGTCTCC ACACCTCCGC TTACATCAAC ACCAATAACG CCATTTAATC  1301  TAAGCGCATC ACCAACATTT TCTGGCGTCA GTCCACCAGC TAACATAAAA  1351  TGTAAGCTTT CGGGGCTCTC TTGCCTTCCA ACCCAGTCAG AAATCGAGTT  1401  CCAATCCAAA AGTTCACCTG TCCCACCTGC TTCTGAATCA AACAAGGGAA  1451  TAAACGAATG AGGTTTCTGT GAAGCTGCAC TGAGTAGTAT GTTGCAGTCT  1501  TTTGGAAATA CGAGTCTTTT AATAACTGGC AAACCGAGGA ACTCTTGGTA  1551  TTCTTGCCAC GACTCATCTC CATGCAGTTG GACGATATCA ATGCCGTAAT  1601  CATTGACCAG AGCCAAAACA TCCTCCTTAG GTTGATTACG AAACACGCCA  1651  ACCAAGTATT TCGGAGTGCC TGAACTATTT TTATATGCTT TTACAAGACT  1701  TGAAATTTTC CTTGCAATAA CCGGGTCAAT TGTTCTCTTT CTATTGGGCA  1751  CACATATAAT ACCCAGCAAG TCAGCATCGG AATCTAGAGC ACATTCTGCG  1801  GCCTCTGTGC TCTGCAAGCC GCAAACTTTC ACCAATGGAC CAGAACTACC  1851  TGTGAAATTA ATAACAGACA TACTCCAAGC TGCCTTTGTG TGCTTAATCA  1901  CGTATACTCA CGTGCTCAAT AGTCACCAAT GCCCTCCCTC TTGGCCCTCT  1951  CCTTTTCTTT TTTCGACCGA ATTAATTCGT AATCATGGTC ATAGCTGTTT  2001  CCTGTGTGAA ATTGTTATCC GCTCACAATT CCACACAACA TACGAGCCGG  2051  AAGCATAAAG TGTAAAGCCT GGGGTGCCTA ATGAGTGAGG TAACTCACAT  2101  TAATTGCGTT GCGCTCACTG CCCGCTTTCC AGTCGGGAAA CCTGTCGTGC  2151  CAGCTGGTTC TTTCCGCCTC AGGACTCTTC CTTTTTCAAT ATTATTGAAG  2201  CATTTATCAG GGTTATTGTC TCATGAGCGG ATACATATTT GAATGTATTT  2251  AGAAAAATAA ACAAATAGGG GTTCCGCGCA CATTTCCCCG AAAAGTGCCA  2301  CCTGACGCGC CCTGTAGCGG CGCATTAAGC GCGGCGGGTG TGGTGGTTAC  2351  GCGCAGCGTG ACCGCTACAC TTGCCAGCGC CCTAGCGCCC GCTCCTTTCG  2401  CTTTCTTCCC TTCCTTTCTC GCCACGTTCG CCGGCTTTCC CCGTCAAGCT  2451  CTAAATCGGG GGCTCCCTTT AGGGTTCCGA TTTAGTGCTT TACGGCACCT  2501  CGACCCCAAA AAACTTGATT AGGGTGATGG TTCACGTAGT GGGCCATCGC  2551  CCTGATAGAC GGTTTTTCGC CCTTTGACGT TGGAGTCCAC GTTCTTTAAT  2601  AGTGGACTCT TGTTCCAAAC TGGAACAACA CTCAACCCTA TCTCGGTCTA  2651  TTCTTTTGAT TTATAAGGGA TTTTGCCGAT TTCGGCCTAT TGGTTAAAAA  2701  ATGAGCTGAT TTAACAAAAA TTTAACGCGA ATTTTAACAA AATATTAACG  2751  CTTACAATTT ACGCGCCTAA ATTATGCGGC CGCATATAAC TGGATTAATG  2801  AATCGGCCAA CGCGCGGGGA GAGGCGGTTT GCGTATTGGG CGCTCTTCCG  2851  CTTCCTCGCT CACTGACTCG CTGCGCTCGG TCGTTCGGCT GCGGCGAGCG  2901  GTATCAGCTC ACTCAAAGGC GGTAATACGG TTATCCACAG AATCAGGGGA  2951  TAACGCAGGA AAGAACATGT GAGCAAAAGG CCAGCAAAAG GCCAGGAACC  3001  GTAAAAAGGC CGCGTTGCTG GCGTTTTTCC ATAGGCTCCG CCCCCCTGAC  3051  GAGCATCACA AAAATCGACG CTCAAGTCAG AGGTGGCGAA ACCCGACAGG  3101  ACTATAAAGA TACCAGGCGT TTCCCCCTGG AAGCTCCCTC GTGCGCTCTC  3151  CTGTTCCGAC CCTGCCGCTT ACCGGATACC TGTCCGCCTT TCTCCCTTCG  3201  GGAAGCGTGG CGCTTTCTCA TAGCTCACGC TGTAGGTATC TCAGTTCGGT  3251  GTAGGTCGTT CGCTCCAAGC TGGGCTGTGT GCACGAACCC CCCGTTCAGC  3301  CCGACCGCTG CGCCTTATCC GGTAACTATC GTCTTGAGTC CAACCCGGTA  3351  AGACACGACT TATCGCCACT GGCAGCAGCC ACTGGTAACA GGATTAGCAG  3401  AGCGAGGTAT GTAGGCGGTG CTACAGAGTT CTTGAAGTGG TGGCCTAACT  3451  ACGGCTACAC TAGAAGGACA GTATTTGGTA TCTGCGCTCT GCTGAAGCCA  3501  GTTACCTTCG GAAAAAGAGT TGGTAGCTCT TGATCCGGCA AACAAACCAC  3551  CGCTGGTAGC GGTGGTTTTT TTGTTTGCAA GCAGCAGATT ACGCGCAGAA  3601  AAAAAGGATC TCAAGAAGAT CCTTTGATCT TTTCTACGGG GTCTGACGCT  3651  CAGTGGAACG AAAACTCACG TTAAGGGATT TTGGTCATGA GATTATCAAA  3701  AAGGATCTTC ACCTAGATCC TTTTAAATTA AAAATGAAGT TTTAAATCAA  3751  TCTAAAGTAT ATATGAGTAA ACTTGGTCTG ACAGTTACCA ATGCTTAATC  3801  AGTGAGGCAC CTATCTCAGC GATCTGTCTA TTTCGTTCAT CCATAGTTGC  3851  CTGACTCCCC GTCGTGTAGA TAACTACGAT ACGGGAGGGC TTACCATCTG  3901  GCCCCAGTGC TGCAATGATA CCGCGAGACC CACGCTCACC GGCTCCAGAT  3951  TTATCAGCAA TAAACCAGCC AGCCGCCTGT GACGGAAGAT CACTTCGCAG  4001  AATAAATAAA TCCTGGTGTC CCTGTTGATA CCGGGAAGCC CTGGGCCAAC  4051  TTTTGGCGAA AATGAGACGT TGATCGGCAC GTAAGAGGTT CCAACTTTCA  4101  CCATAATGAA ATAAGATCAC TACCGGGCGT ATTTTTTGAG TTGTCGAGAT  4151  TTTCAGGAGC TAAGGAAGCT AAAATGGAGA AAAAAATCAC TGGATATACC  4201  ACCGTTGATA TATCCCAATG GCATCGTAAA GAACATTTTG AGGCATTTCA  4251  GTCAGTTGCT CAATGTACCT ATAACCAGAC CGTTCAGCTG GATATTACGG  4301  CCTTTTTAAA GACCGTAAAG AAAAATAAGC ACAAGTTTTA TCCGGCCTTT  4351  ATTCACATTC TTGCCCGCCT GATGAATGCT CATCCGGAAT TACGTATGGC  4401  AATGAAAGAC GGTGAGCTGG TGATATGGGA TAGTGTTCAC CCTTGTTACA  4451  CCGTTTTCCA TGAGCAAACT GAAACGTTTT CATCGCTCTG GAGTGAATAC  4501  CACGACGATT TCCGGCAGTT TCTACACATA TATTCGCAAG ATGTGGCGTG  4551  TTACGGTGAA AACCTGGCCT ATTTCCCTAA AGGGTTTATT GAGAATATGT  4601  TTTTCGTCTC AGCCAATCCC TGGGTGAGTT TCACCAGTTT TGATTTAAAC  4651  GTGGCCAATA TGGACAACTT CTTCGCCCCG TTTTCACCAT GGGCAAATAT  4701  TATACGCAAG GCGACAAGGT GCTGATGCCG CTGGCGATTC AGGTTCATCA  4751  TGCCGTTTGT GATGGCTTCC ATGTCGGCAG AATGCTTAAT GAATTACAAC  4801  AGTACTGCGA TGAGTGGCAG GGCGGGGCGT AATTTTTTTA AGGCAGTTAT  4851  TGGTGCCCTT AAACGCCTGG TTGCTACGCC TGAATAAGTG ATAATAAGCG  4901  GATGAATGGC AGAAATTCGA AAGCAAATTC GACCCGGTCG TCGGTTCAGG  4951  GCAGGGTCGT TAAATAGCCG CTTATGTCTA TTGCTGGTTT ACCGGTTTAT  5001  TGACTACCGG AAGCAGTGTG ACCGTGTGCT TCTCAAATGC CTGAGGCCAG  5051  TTTGCTCAGG CTCTCCCCGT GGAGGCTAAG AAACCATTAT TATCATGACA  5101  TTAACCTATA AAAATAGGCG TATCACGAGG CCCTTTCGTC TCGCGCGTTT  5151  CGGTGATGAC GGTGAAAACC TCTGACACAT GCAGCTCCCG GAGACGGTCA  5201  CAGCTTGTCT GTAAGCGGAT GCCGGGAGCA GACAAGCCCG TCAGGGCGCG  5251  TCAGCGGGTG TTGGCGGGTG TCGGGGCTGG CTTAACTATG CGGCATCAGA  5301  GCAGATTGTA CTGAGAGTGC ACCATAACGC ATTTAAGCAT AAACACGCAC  5351  TATGCCGTTC TTCTCATGTA TATATATATA CAGGCAACAC GCAGATATAG  5401  GTGCGACGTG AACAGTGAGC TGTATGTGCG CAGCTCGCGT TGCATTTTCG  5451  GAAGCGCTCG TTTTCGGAAA CGCTTTGAAG TTCCTATTCC GAAGTTCCTA  5501  TTCTCTAGCT AGAAAGTATA GGAACTTCAG AGCGCTTTTG AAAACCAAAA  5551  GCGCTCTGAA GACGCACTTT CAAAAAACCA AAAACGCACC GGACTGTAAC  5601  GAGCTACTAA AATATTGCGA ATACCGCTTC CACAAACATT GCTCAAAAGT  5651  ATCTCTTTGC TATATATCTC TGTGCTATAT CCCTATATAA CCTACCCATC  5701  CACCTTTCGC TCCTTGAACT TGCATCTAAA CTCGACCTCT ACATTTTTTA  5751  TGTTTATCTC TAGTATTACT CTTTAGACAA AAAAATTGTA GTAAGAACTA  5801  TTCATAGAGT GAATCGAAAA CAATACGAAA ATGTAAACAT TTCCTATACG  5851  TAGTATATAG AGACAAAATA GAAGAAACCG TTCATAATTT TCTGACCAAT  5901  GAAGAATCAT CAACGCTATC ACTTTCTGTT CACAAAGTAT GCGCAATCCA  5951  CATCGGTATA GAATATAATC GGGGATGCCT TTATCTTGAA AAAATGCACC  6001  CGCAGCTTCG CTAGTAATCA GTAAACGCGG GAAGTGGAGT CAGGCTTTTT  6051  TTATGGAAGA GAAAATAGAC ACCAAAGTAG CCTTCTTCTA ACCTTAACGG  6101  ACCTACAGTG CAAAAAGTTA TCAAGAGACT GCATTATAGA GCGCACAAAG  6151  GAGAAAAAAA GTAATCTAAG ATGCTTTGTT AGAAAAATAG CGCTCTCGGG  6201  ATGCATTTTT GTAGAACAAA AAAGAAGTAT AGATTCTTTG TTGGTAAAAT  6251  AGCGCTCTCG CGTTGCATTT CTGTTCTGTA AAAATGCAGC TCAGATTCTT  6301  TGTTTGAAAA ATTAGCGCTC TCGCGTTGCA TTTTTGTTTT ACAAAAATGA  6351  AGCACAGATT CTTCGTTGGT AAAATAGCGC TTTCGCGTTG CATTTCTGTT  6401  CTGTAAAAAT GCAGCTCAGA TTCTTTGTTT GAAAAATTAG CGCTCTCGCG  6451  TTGCATTTTT GTTCTACAAA ATGAAGCACA GATGCTTCGT T