pET-33b(+)
pET-33b(+)
编号 | 名称 |
北京华越洋VECT-590 | pET-33b(+) |
pET33b载体基本信息
别名: | pET33b, pET 33b |
质粒类型: | 大肠杆菌蛋白表达 |
表达水平: | 高 |
克隆方法: | 多克隆位点,限制性内切酶 |
载体大小: | 5383 bp |
5' 测序引物: | T7 |
5' 测序引物序列: | T7: 5'-TAATACGACTCACTATAGGG-3' |
载体标签: | His (中间和C端), N-Thrombin |
载体抗性: | Kanamycin |
备注: | Production of target proteins suitable for site-specific 32P-labeling; Nterm thrombin cleavage site; Nterm enterokinase cleavage site |
稳定性: | 瞬时表达 Transient |
组成型: | 组成型 Constitutive |
病毒/非病毒: | 非病毒 |
pET33b载体质粒图谱和多克隆位点信息
Feature Name | Start | End |
T7_terminator | 129 | 1 |
T7_Terminal_primer | 69 | 87 |
T7_leader | 238 | 206 |
Xpress_fwd_primer | 239 | 221 |
T7_transl_en_RBS | 337 | 321 |
lacO | 382 | 355 |
T7_promoter | 400 | 382 |
tet (300 - 563) | 432 | 695 |
pBRrevBam_primer | 503 | 484 |
lacI | 778 | 1869 |
tet (576 - 851) | 1928 | 2203 |
ROP | 2678 | 2869 |
pGEX_3_primer | 2885 | 2863 |
pBR322_origin | 3903 | 3284 |
KanR2 | 4009 | 4824 |
ORF | Start | End |
ORF frame 3 | 537 | 1037 |
ORF frame 1 | 910 | 1869 |
ORF frame 1 | 4009 | 4824 |
Enzyme Name | Cut |
XhoI | 158 |
NotI | 166 |
EagI | 166 |
HindIII | 173 |
SalI | 179 |
SacI | 190 |
EcoRI | 192 |
BamHI | 198 |
NheI | 230 |
NdeI | 237 |
NcoI | 310 |
XbaI | 349 |
BglII | 415 |
BclI | 1151 |
ApaI | 1348 |
EcoRV | 1587 |
HpaI | 1643 |
FspI | 2219 |
NruI | 4097 |
ClaI | 4131 |
XmaI | 4312 |
SmaI | 4314 |
pET33b载体简介
The pET-33b(+) vector is derived from pET-28b(+) and carries a 15bp sequence encoding the protein kinase A (PKA) site RRASV, located between the thrombin cleavage and Nde I sites (1). Proteins expressed in pET-33b(+) can be easily purified by metal chelation chromatography (via either N- or C-terminal His•Tag® sequences) and efficiently labeled with 32P- or 33P-gATP and the catalytic subunit of cAMP-dependent protein kinase from heart muscle. Labeled proteins can be used as direct probes in protein-protein interaction studies. Unique sites are shown on the circle map. Note that the sequence is numbered by the pBR322 convention, so the T7 expression region is reversed on the circle map. The cloning/ expression region of the coding strand transcribed by T7 RNA polymerase is shown below. The f1 origin is oriented so that infection with helper phage will produce virions containing single-stranded DNA that corresponds to the coding strand. Therefore, single-stranded sequencing should be performed using the T7 terminator primer.
pET33b载体序列
ORIGIN
1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtgcggcc gcaagcttgt
181 cgacggagct cgaattcgga tcccgaccca tttgctgtcc accagtcatg ctagccatat
241 gaacagatgc acgacggctg ccgcgcggca ccaggccgct gctgtgatga tgatgatgat
301 ggctgctgcc catggtatat ctccttctta aagttaaaca aaattatttc tagaggggaa
361 ttgttatccg ctcacaattc ccctatagtg agtcgtatta atttcgcggg atcgagatct
421 cgatcctcta cgccggacgc atcgtggccg gcatcaccgg cgccacaggt gcggttgctg
481 gcgcctatat cgccgacatc accgatgggg aagatcgggc tcgccacttc gggctcatga
541 gcgcttgttt cggcgtgggt atggtggcag gccccgtggc cgggggactg ttgggcgcca
601 tctccttgca tgcaccattc cttgcggcgg cggtgctcaa cggcctcaac ctactactgg
661 gctgcttcct aatgcaggag tcgcataagg gagagcgtcg agatcccgga caccatcgaa
721 tggcgcaaaa cctttcgcgg tatggcatga tagcgcccgg aagagagtca attcagggtg
781 gtgaatgtga aaccagtaac gttatacgat gtcgcagagt atgccggtgt ctcttatcag
841 accgtttccc gcgtggtgaa ccaggccagc cacgtttctg cgaaaacgcg ggaaaaagtg
901 gaagcggcga tggcggagct gaattacatt cccaaccgcg tggcacaaca actggcgggc
961 aaacagtcgt tgctgattgg cgttgccacc tccagtctgg ccctgcacgc gccgtcgcaa
1021 attgtcgcgg cgattaaatc tcgcgccgat caactgggtg ccagcgtggt ggtgtcgatg
1081 gtagaacgaa gcggcgtcga agcctgtaaa gcggcggtgc acaatcttct cgcgcaacgc
1141 gtcagtgggc tgatcattaa ctatccgctg gatgaccagg atgccattgc tgtggaagct
1201 gcctgcacta atgttccggc gttatttctt gatgtctctg accagacacc catcaacagt
1261 attattttct cccatgaaga cggtacgcga ctgggcgtgg agcatctggt cgcattgggt
1321 caccagcaaa tcgcgctgtt agcgggccca ttaagttctg tctcggcgcg tctgcgtctg
1381 gctggctggc ataaatatct cactcgcaat caaattcagc cgatagcgga acgggaaggc
1441 gactggagtg ccatgtccgg ttttcaacaa accatgcaaa tgctgaatga gggcatcgtt
1501 cccactgcga tgctggttgc caacgatcag atggcgctgg gcgcaatgcg cgccattacc
1561 gagtccgggc tgcgcgttgg tgcggatatc tcggtagtgg gatacgacga taccgaagac
1621 agctcatgtt atatcccgcc gttaaccacc atcaaacagg attttcgcct gctggggcaa
1681 accagcgtgg accgcttgct gcaactctct cagggccagg cggtgaaggg caatcagctg
1741 ttgcccgtct cactggtgaa aagaaaaacc accctggcgc ccaatacgca aaccgcctct
1801 ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc
1861 gggcagtgag cgcaacgcaa ttaatgtaag ttagctcact cattaggcac cgggatctcg
1921 accgatgccc ttgagagcct tcaacccagt cagctccttc cggtgggcgc ggggcatgac
1981 tatcgtcgcc gcacttatga ctgtcttctt tatcatgcaa ctcgtaggac aggtgccggc
2041 agcgctctgg gtcattttcg gcgaggaccg ctttcgctgg agcgcgacga tgatcggcct
2101 gtcgcttgcg gtattcggaa tcttgcacgc cctcgctcaa gccttcgtca ctggtcccgc
2161 caccaaacgt ttcggcgaga agcaggccat tatcgccggc atggcggccc cacgggtgcg
2221 catgatcgtg ctcctgtcgt tgaggacccg gctaggctgg cggggttgcc ttactggtta
2281 gcagaatgaa tcaccgatac gcgagcgaac gtgaagcgac tgctgctgca aaacgtctgc
2341 gacctgagca acaacatgaa tggtcttcgg tttccgtgtt tcgtaaagtc tggaaacgcg
2401 gaagtcagcg ccctgcacca ttatgttccg gatctgcatc gcaggatgct gctggctacc
2461 ctgtggaaca cctacatctg tattaacgaa gcgctggcat tgaccctgag tgatttttct
2521 ctggtcccgc cgcatccata ccgccagttg tttaccctca caacgttcca gtaaccgggc
2581 atgttcatca tcagtaaccc gtatcgtgag catcctctct cgtttcatcg gtatcattac
2641 ccccatgaac agaaatcccc cttacacgga ggcatcagtg accaaacagg aaaaaaccgc
2701 ccttaacatg gcccgcttta tcagaagcca gacattaacg cttctggaga aactcaacga
2761 gctggacgcg gatgaacagg cagacatctg tgaatcgctt cacgaccacg ctgatgagct
2821 ttaccgcagc tgcctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct
2881 cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg
2941 cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg acccagtcac gtagcgatag
3001 cggagtgtat actggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat
3061 atatgcggtg tgaaataccg cacagatgcg taaggagaaa ataccgcatc aggcgctctt
3121 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
3181 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
3241 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
3301 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
3361 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
3421 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
3481 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
3541 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
3601 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
3661 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
3721 actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct
3781 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
3841 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
3901 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
3961 tgaacaataa aactgtctgc ttacataaac agtaatacaa ggggtgttat gagccatatt
4021 caacgggaaa cgtcttgctc taggccgcga ttaaattcca acatggatgc tgatttatat
4081 gggtataaat gggctcgcga taatgtcggg caatcaggtg cgacaatcta tcgattgtat
4141 gggaagcccg atgcgccaga gttgtttctg aaacatggca aaggtagcgt tgccaatgat
4201 gttacagatg agatggtcag actaaactgg ctgacggaat ttatgcctct tccgaccatc
4261 aagcatttta tccgtactcc tgatgatgca tggttactca ccactgcgat ccccgggaaa
4321 acagcattcc aggtattaga agaatatcct gattcaggtg aaaatattgt tgatgcgctg
4381 gcagtgttcc tgcgccggtt gcattcgatt cctgtttgta attgtccttt taacagcgat
4441 cgcgtatttc gtctcgctca ggcgcaatca cgaatgaata acggtttggt tgatgcgagt
4501 gattttgatg acgagcgtaa tggctggcct gttgaacaag tctggaaaga aatgcataaa
4561 cttttgccat tctcaccgga ttcagtcgtc actcatggtg atttctcact tgataacctt
4621 atttttgacg aggggaaatt aataggttgt attgatgttg gacgagtcgg aatcgcagac
4681 cgataccagg atcttgccat cctatggaac tgcctcggtg agttttctcc ttcattacag
4741 aaacggcttt ttcaaaaata tggtattgat aatcctgata tgaataaatt gcagtttcat
4801 ttgatgctcg atgagttttt ctaagaatta attcatgagc ggatacatat ttgaatgtat
4861 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgaaat
4921 tgtaaacgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt
4981 taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg
5041 gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt
5101 caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc
5161 aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg
5221 atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa
5281 aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc
5341 cgccgcgctt aatgcgccgc tacagggcgc gtcccattcg cca
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