| 出品公司: | Invitrogen |
|---|---|
| 载体名称: | pYES2-CT, pYES2/CT |
| 质粒类型: | 酿酒酵母表达载体 |
| 表达水平: | 高拷贝 |
| 诱导方法: | 半乳糖 |
| 启动子: | GAL1 |
| 克隆方法: | 多克隆位点，限制性内切酶 |
| 载体大小: | 5963 bp |
| 5' 测序引物及序列: | GAL1-F: AATATACCTCTATACTTTAACGTC |
| 3' 测序引物及序列: | CYC1-R: GCGTGAATGTAAGCGTGAC |
| 载体标签: | C-His, C-V5 |
| 载体抗性: | 氨苄青霉素 |
| 筛选标记: | URA3报告基因 |
| 克隆菌株: | TOP10, E.coli |
| 表达菌株: | INVSc1, Saccharomyces cerevisiae |
| 备注: |
酿酒酵母表达载体pYES2/CT含有URA3报告基因； GAL1启动子驱动目的基因高水平表达； 半乳糖诱导目基因表达，葡萄糖阻遏目的基因表达，二者起到分子开关作用。 |
| 产品目录号: | V8251-20 |
| 稳定性: | 稳表达 |
| 组成型/诱导型: | 诱导型 |
| 病毒/非病毒: | 非病毒 |
买家导航
pYES2/CT载体质粒基本信息
pYES2/CT质粒图谱载体图谱和pYES2/CT载体序列质粒序列多克隆位点信息
pYES2/CT质粒载体简介
pYES2/CT酿酒酵母表达载体
pYES2/CT载体大小为6.0 kb左右，用于重组蛋白在酿酒酵母细胞中的诱导型表达。诱导剂为半乳糖。重组蛋白的C-端融合了6XHis标签，用于重组蛋白的纯化和检测。
pYES2/CT载体含有以下元件：
Yeast GAL1 promoter for high level inducible protein expression in yeast by galactose and repression by glucose (Giniger et al., 1985; West et al., 1984)
Multiple cloning site (MCS) with 8 or 9 unique sites (plus two BstX I sites) to facilitate in-frame cloning with the C-terminal peptide
C-terminal peptide encoding the V5 epitope and a polyhistidine (6xHis) tag for detection and purification of your recombinant fusion protein
2μ origin for episomal maintenance and high copy replication or CEN6/ARSH4 sequence for non-integrative centromeric maintenance and low copy replication (pYC2/CT)
URA3 auxotrophic marker for selection of yeast transformants
Ampicillin resistance gene for selection in E. coli
使用pYES2/CT载体表达目的基因的实验流程如下：
1 Consult the multiple cloning site described on page 7 to determine a strategy to clone your gene in frame with the C-terminal peptide.
2 Ligate your insert into the appropriate vector and transform into E. coli. Select transformants on LB plates containing 50 to 100 μg/mL ampicillin.
3 Analyze your transformants for the presence of insert by restriction digestion.
4 Select a transformant with the correct restriction pattern and sequence to confirm that your gene is cloned in frame with the C-terminal peptide.
5 Transform your construct into competent INVSc1 cells and select for the appropriate amino acid prototrophy.
6 Test for expression of your recombinant protein by western blot analysis or functional assay.
7 Use metal-chelating resin such as ProBond to purify your recombinant protein.
pYES2/CT质粒序列载体序列
LOCUS pYES2/CT 5963 bp DNA circular SYN
DEFINITION pYES2/CT
ACCESSION
KEYWORDS
SOURCE
ORGANISM other sequences; artificial sequences; vectors.
COMMENT This file is created by Vector NTI
http://www.biofeng.com/
COMMENT VNTAUTHORNAME|biofeng.com|
FEATURES Location/Qualifiers
source 1..5963
/organism="pYES2/CT"
/mol_type="other DNA"
promoter 1..451
/label="GAL1_promoter"
misc_feature 414..437
/label="GAL1_primer"
promoter 475..493
/label="T7_promoter"
terminator 716..961
/label="CYC1_terminator"
misc_feature complement(725..743)
/label="pYESTrp_rev_primer"
misc_feature complement(725..743)
/label="CYC1_primer"
rep_origin complement(1189..1808)
/label="pBR322_origin"
CDS complement(1963..2823)
/label="ORF frame 3"
CDS complement(2919..3722)
/label="ORF frame 1"
gene complement(2922..3722)
/label="URA3"
/gene="URA3"
CDS 3211..3687
/label="ORF frame 1"
promoter complement(3724..3949)
/label="URA3_promoter"
rep_origin 3952..5423
/label="2micron_origin"
ORIGIN
1 ACGGATTAGA AGCCGCCGAG CGGGTGACAG CCCTCCGAAG GAAGACTCTC CTCCGTGCGT
61 CCTCGTCTTC ACCGGTCGCG TTCCTGAAAC GCAGATGTGC CTCGCGCCGC ACTGCTCCGA
121 ACAATAAAGA TTCTACAATA CTAGCTTTTA TGGTTATGAA GAGGAAAAAT TGGCAGTAAC
181 CTGGCCCCAC AAACCTTCAA ATGAACGAAT CAAATTAACA ACCATAGGAT GATAATGCGA
241 TTAGTTTTTT AGCCTTATTT CTGGGGTAAT TAATCAGCGA AGCGATGATT TTTGATCTAT
301 TAACAGATAT ATAAATGCAA AAACTGCATA ACCACTTTAA CTAATACTTT CAACATTTTC
361 GGTTTGTATT ACTTCTTATT CAAATGTAAT AAAAGTATCA ACAAAAAATT GTTAATATAC
421 CTCTATACTT TAACGTCAAG GAGAAAAAAC CCCGGATCGG ACTACTAGCA GCTGTAATAC
481 GACTCACTAT AGGGAATATT AAGCTTGGTA CCGAGCTCGG ATCCACTAGT AACGGCCGCC
541 AGTGTGCTGG AATTCTGCAG ATATCCAGCA CAGTGGCGGC CGCTCGAGTC TAGAGGGCCC
601 TTCGAAGGTA AGCCTATCCC TAACCCTCTC CTCGGTCTCG ATTCTACGCG TACCGGTCAT
661 CATCACCATC ACCATTGAGT TTAAACCCGC TGATCCTAGA GGGCCGCATC ATGTAATTAG
721 TTATGTCACG CTTACATTCA CGCCCTCCCC CCACATCCGC TCTAACCGAA AAGGAAGGAG
781 TTAGACAACC TGAAGTCTAG GTCCCTATTT ATTTTTTTAT AGTTATGTTA GTATTAAGAA
841 CGTTATTTAT ATTTCAAATT TTTCTTTTTT TTCTGTACAG ACGCGTGTAC GCATGTAACA
901 TTATACTGAA AACCTTGCTT GAGAAGGTTT TGGGACGCTC GAAGGCTTTA ATTTGCAAGC
961 TGCGGCCCTG CATTAATGAA TCGGCCAACG CGCGGGGAGA GGCGGTTTGC GTATTGGGCG
1021 CTCTTCCGCT TCCTCGCTCA CTGACTCGCT GCGCTCGGTC GTTCGGCTGC GGCGAGCGGT
1081 ATCAGCTCAC TCAAAGGCGG TAATACGGTT ATCCACAGAA TCAGGGGATA ACGCAGGAAA
1141 GAACATGTGA GCAAAAGGCC AGCAAAAGCC CAGGAACCGT AAAAAGGCCG CGTTGCTGGC
1201 GTTTTTCCAT AGGCTCCGCC CCCCTGACGA GCATCACAAA AATCGACGCT CAAGTCAGAG
1261 GTGGCGAAAC CCGACAGGAC TATAAAGATA CCAGGCGTTT CCCCCTGGAA GCTCCCTCGT
1321 GCGCTCTCCT GTTCCGACCC TGCCGCTTAC CGGATACCTG TCCGCCTTTC TCCCTTCGGG
1381 AAGCGTGGCG CTTTCTCATA GCTCACGCTG TAGGTATCTC AGTTCGGTGT AGGTCGTTCG
1441 CTCCAAGCTG GGCTGTGTGC ACGAACCCCC CGTTCAGCCC GACCGCTGCG CCTTATCCGG
1501 TAACTATCGT CTTGAGTCCA ACCCGGTAAG ACACGACTTA TCGCCACTGG CAGCAGCCAC
1561 TGGTAACAGG ATTAGCAGAG CGAGGTATGT AGGCGGTGCT ACAGAGTTCT TGAAGTGGTG
1621 GCCTAACTAC GGCTACACTA GAAGGACAGT ATTTGGTATC TGCGCTCTGC TGAAGCCAGT
1681 TACCTTCGGA AAAAGAGTTG GTAGCTCTTG ATCCGGCAAA CAAACCACCG CTGGTAGCGG
1741 TGGTTTTTTT GTTTGCAAGC AGCAGATTAC GCGCAGAAAA AAAGGATCTC AAGAAGATCC
1801 TTTGATCTTT TCTACGGGGT CTGACGCTCA GTGGAACGAA AACTCACGTT AAGGGATTTT
1861 GGTCATGAGA TTATCAAAAA GGATCTTCAC CTAGATCCTT TTAAATTAAA AATGAAGTTT
1921 TAAATCAATC TAAAGTATAT ATGAGTAAAC TTGGTCTGAC AGTTACCAAT GCTTAATCAG
1981 TGAGGCACCT ATCTCAGCGA TCTGTCTATT TCGTTCATCC ATAGTTGCCT GACTCCCCGT
2041 CGTGTAGATA ACTACGATAC GGGAGCGCTT ACCATCTGGC CCCAGTGCTG CAATGATACC
2101 GCGAGACCCA CGCTCACCGG CTCCAGATTT ATCAGCAATA AACCAGCCAG CCGGAAGGGC
2161 CGAGCGCAGA AGTGGTCCTG CAACTTTATC CGCCTCCATC CAGTCTATTA ATTGTTGCCG
2221 GGAAGCTAGA GTAAGTAGTT CGCCAGTTAA TAGTTTGCGC AACGTTGTTG GCATTGCTAC
2281 AGGCATCGTG GTGTCACTCT CGTCGTTTGG TATGGCTTCA TTCAGCTCCG GTTCCCAACG
2341 ATCAAGGCGA GTTACATGAT CCCCCATGTT GTGCAAAAAA GCGGTTAGCT CCTTCGGTCC
2401 TCCGATCGTT GTCAGAAGTA AGTTGGCCGC AGTGTTATCA CTCATGGTTA TGGCAGCACT
2461 GCATAATTCT CTTACTGTCA TGCCATCCGT AAGATGCTTT TCTGTGACTG GTGAGTACTC
2521 AACCAAGTCA TTCTGAGAAT AGTGTATGCG GCGACCGAGT TGCTCTTGCC CGGCGTCAAT
2581 ACGGGATAAT AGTGTATCAC ATAGCAGAAC TTTAAAAGTG CTCATCATTG GAAAACGTTC
2641 TTCGGGGCGA AAACTCTCAA GGATCTTACC GCTGTTGAGA TCCAGTTCGA TGTAACCCAC
2701 TCGTGCACCC AACTGATCTT CAGCATCTTT TACTTTCACC AGCGTTTCTG GGTGAGCAAA
2761 AACAGGAAGG CAAAATGCCG CAAAAAAGGG AATAAGGGCG ACACGGAAAT GTTGAATACT
2821 CATACTCTTC CTTTTTCAAT GGGTAATAAC TGATATAATT AAATTGAAGC TCTAATTTGT
2881 GAGTTTAGTA TACATGCATT TACTTATAAT ACAGTTTTTT AGTTTTGCTG GCCGCATCTT
2941 CTCAAATATG CTTCCCAGCC TGCTTTTCTG TAACGTTCAC CCTCTACCTT AGCATCCCTT
3001 CCCTTTGCAA ATAGTCCTCT TCCAACAATA ATAATGTCAG ATCCTGTAGA GACCACATCA
3061 TCCACGGTTC TATACTGTTG ACCCAATGCG TCTCCCTTGT CATCTAAACC CACACCGGGT
3121 GTCATAATCA ACCAATCGTA ACCTTCATCT CTTCCACCCA TGTCTCTTTG AGCAATAAAG
3181 CCGATAACAA AATCTTTGTC GCTCTTCGCA ATGTCAACAG TACCCTTAGT ATATTCTCCA
3241 GTAGATAGGG AGCCCTTGCA TGACAATTCT GCTAACATCA AAAGGCCTCT AGGTTCCTTT
3301 GTTACTTCTT CTGCCGCCTG CTTCAAACCG CTAACAATAC CTGGGCCCAC CACACCGTGT
3361 GCATTCGTAA TGTCTGCCCA TTCTGCTATT CTGTATACAC CCGCAGAGTA CTGCAATTTG
3421 ACTGTATTAC CAATGTCAGC AAATTTTCTG TCTTCGAAGA GTAAAAAATT GTACTTGGCG
3481 GATAATGCCT TTAGCGGCTT AACTGTGCCC TCCATGGAAA AATCAGTCAA GATATCCACA
3541 TGTGTTTTTA GTAAACAAAT TTTGGGACCT AATGCTTCAA CTAACTCCAG TAATTCCTTG
3601 GTGGTACGAA CATCCAATGA AGCACACAAG TTTGTTTGCT TTTCGTGCAT GATATTAAAT
3661 AGCTTGGCAG CAACAGGACT AGGATGAGTA GCAGCACGTT CCTTATATGT AGCTTTCGAC
3721 ATGATTTATC TTCGTTTCCT GCAGGTTTTT GTTCTGTGCA GTTGGGTTAA GAATACTGGG
3781 CAATTTCATG TTTCTTCAAC ACTACATATG CGTATATATA CCAATCTAAG TCTGTGCTCC
3841 TTCCTTCGTT CTTCCTTCTG TTCGGAGATT ACCGAATCAA AAAAATTTCA AAGAAACCGA
3901 AATCAAAAAA AAGAATAAAA AAAAAATGAT GAATTGAATT GAAAAGCTAG CTTATCGATG
3961 ATAAGCTGTC AAAGATGAGA ATTAATTCCA CGGACTATAG ACTATACTAG ATACTCCGTC
4021 TACTGTACGA TACACTTCCG CTCAGGTCCT TGTCCTTTAA CGAGGCCTTA CCACTCTTTT
4081 GTTACTCTAT TGATCCAGCT CAGCAAAGGC AGTGTGATCT AAGATTCTAT CTTCGCGATG
4141 TAGTAAAACT AGCTAGACCG AGAAAGAGAC TAGAAATGCA AAAGGCACTT CTACAATGGC
4201 TGCCATCATT ATTATCCGAT GTGACGCTGC AGCTTCTCAA TGATATTCGA ATACGCTTTG
4261 AGGAGATACA GCCTAATATC CGACAAACTG TTTTACAGAT TTACGATCGT ACTTGTTACC
4321 CATCATTGAA TTTTGAACAT CCGAACCTGG GAGTTTTCCC TGAAACAGAT AGTATATTTG
4381 AACCTGTATA ATAATATATA GTCTAGCGCT TTACGGAAGA CAATGTATGT ATTTCGGTTC
4441 CTGGAGAAAC TATTGCATCT ATTGCATAGG TAATCTTGCA CGTCGCATCC CCGGTTCATT
4501 TTCTGCGTTT CCATCTTGCA CTTCAATAGC ATATCTTTGT TAACGAAGCA TCTGTGCTTC
4561 ATTTTGTAGA ACAAAAATGC AACGCGAGAG CGCTAATTTT TCAAACAAAG AATCTGAGCT
4621 GCATTTTTAC AGAACAGAAA TGCAACGCGA AAGCGCTATT TTACCAACGA AGAATCTGTG
4681 CTTCATTTTT GTAAAACAAA AATGCAACGC GACGAGAGCG CTAATTTTTC AAACAAAGAA
4741 TCTGAGCTGC ATTTTTACAG AACAGAAATG CAACGCGAGA GCGCTATTTT ACCAACAAAG
4801 AATCTATACT TCTTTTTTGT TCTACAAAAA TGCATCCCGA GAGCGCTATT TTTCTAACAA
4861 AGCATCTTAG ATTACTTTTT TTCTCCTTTG TGCGCTCTAT AATGCAGTCT CTTGATAACT
4921 TTTTGCACTG TAGGTCCGTT AAGGTTAGAA GAAGGCTACT TTGGTGTCTA TTTTCTCTTC
4981 CATAAAAAAA GCCTGACTCC ACTTCCCGCG TTTACTGATT ACTAGCGAAG CTGCGGGTGC
5041 ATTTTTTCAA GATAAAGGCA TCCCCGATTA TATTCTATAC CGATGTGGAT TGCGCATACT
5101 TTGTGAACAG AAAGTGATAG CGTTGATGAT TCTTCATTGG TCAGAAAATT ATGAACGGTT
5161 TCTTCTATTT TGTCTCTATA TACTACGTAT AGGAAATGTT TACATTTTCG TATTGTTTTC
5221 GATTCACTCT ATGAATAGTT CTTACTACAA TTTTTTTGTC TAAAGAGTAA TACTAGAGAT
5281 AAACATAAAA AATGTAGAGG TCGAGTTTAG ATGCAAGTTC AAGGAGCGAA AGGTGGATGG
5341 GTAGGTTATA TAGGGATATA GCACAGAGAT ATATAGCAAA GAGATACTTT TGAGCAATGT
5401 TTGTGGAAGC GGTATTCGCA ATGGGAAGCT CCACCCCGGT TGATAATCAG AAAAGCCCCA
5461 AAAACAGGAA GATTGTATAA GCAAATATTT AAATTGTAAA CGTTAATATT TTGTTAAAAT
5521 TCGCGTTAAA TTTTTGTTAA ATCAGCTCAT TTTTTAACGA ATAGCCCGAA ATCGGCAAAA
5581 TCCCTTATAA ATCAAAAGAA TAGACCGAGA TAGGGTTGAG TGTTGTTCCA GTTTCCAACA
5641 AGAGTCCACT ATTAAAGAAC GTGGACTCCA ACGTCAAAGG GCGAAAAAGG GTCTATCAGG
5701 GCGATGGCCC ACTACGTGAA CCATCACCCT AATCAAGTTT TTTGGGGTCG AGGTGCCGTA
5761 AAGCAGTAAA TCGGAAGGGT AAACGGATGC CCCCATTTAG AGCTTGACGG GGAAAGCCGG
5821 CGAACGTGGC GAGAAAGGAA GGGAAGAAAG CGAAAGGAGC GGGGGCTAGG GCGGTGGGAA
5881 GTGTAGGGGT CACGCTGGGC GTAACCACCA CACCCGCCGC GCTTAATGGG GCGCTACAGG
5941 GCGCGTGGGG ATGATCCACT AGT
//
