pET-35b(+)

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pET35b载体基本信息

出品公司: Novagen
载体名称: pET-35b(+)
质粒类型: 大肠杆菌表达
启动子: T7
载体大小: 5927bp
5' 测序引物及序列: T7:TAATACGACTCACTATAGGG
3' 测序引物及序列: T7-ter:TGCTAGTTATTGCTCAGCGG
载体标签: N-CBDclos,C-6×His
载体抗性: 卡那霉素
克隆菌株: DH5α 等
表达宿主: BL21(DE3)
培养条件: 37℃,有氧,LB
诱导方式: IPTG或乳糖及其类似物

pET35b载体质粒图谱和多克隆位点信息

pET-35b载体图谱
pET35b质粒图谱

pET35b载体简介

pET-35b(+)(Cat. No. 70103-3) 号载体是为表达cbdclos融合蛋白而设计的有多种克隆地点和策略可供选择。pCR扩增的快速定向克隆pET-35xa/lic载体试剂盒(Cat. No. 70115-3),包含线性化pET-35B(+)已准备好,可使用适当制备的插件进行退火。当用xa/lic克隆时产生的cbdclos•tag™融合蛋白可以在编码的载体上精确地切割使用因子xa连接。独特的地点显示在圆形地图上。请注意,序列是按PBR322约定编号的,因此T7表达式区域在圆映射上反转。这个 由t7 rna聚合酶转录的编码链的克隆/表达区域如下所示。f1基因的起源是定向的,因此与辅助噬菌体的感染将产生含有单个与编码链相对应的链状DNA。因此,单链测序应该使用T7终端底漆(Cat. No. 69337-3)。

pET35b载体序列

LOCUS       pET-35b                5927 bp ds-DNA     circular SYN 07-8-2015
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    Untitled
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5927)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported 2015-8-7  from MLCC
            http://www.miaolingbio.com
FEATURES             Location/Qualifiers
     source          1..5927
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     source          860..882
                     /organism="Enterobacteria phage T7"
                     /mol_type="genomic DNA"
                     /db_xref="taxon:10760"
     terminator      26..73
                     /note="T7 terminator"
                     /note="transcription terminator for bacteriophage T7 RNA
                     polymerase"
     CDS             complement(150..173)
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /note="8xHis"
                     /translation="HHHHHHHH"
     CDS             complement(249..260)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /note="Factor Xa site"
                     /translation="IEGR"
     CDS             complement(291..335)
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic
                     ribonuclease A"
                     /note="S-Tag"
                     /translation="KETAAAKFERQHMDS"
     CDS             complement(351..368)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /note="thrombin site"
                     /translation="LVPRGS"
     RBS             860..882
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    897..921
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="lac operator"
                     /note="The lac repressor binds to the lac operator to
                     inhibit transcription in E. coli. This inhibition can be
                     relieved by adding lactose or
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(922..940)
                     /note="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        1253..1330
                     /gene="lacI"
                     /note="lacI promoter"
     CDS             1331..2413
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /note="lacI"
                     /note="The lac repressor binds to the lac operator to
                     inhibit transcription in E. coli. This inhibition can be
                     relieved by adding lactose or
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     CDS             3222..3413
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /note="rop"
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     misc_feature    3515..3657
                     /note="bom"
                     /note="basis of mobility region from pBR322"
     rep_origin      complement(3843..4431)
                     /direction=LEFT
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     CDS             4553..5368
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /note="KanR"
                     /note="confers resistance to kanamycin in bacteria or G418
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      complement(5461..5916)
                     /direction=LEFT
                     /note="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgtttagcag cctaggtatt aatcaattag tggtggtggt ggtggtggtg gtgctcgagt
      181 gcggccgcaa gcttgtcgac ggagctcgaa ttcggatccg atatcgccat ggccagagga
      241 gagttagagc gaccctcaat accggagcca ccaccggtac ccagatctgg gctgtccatg
      301 tgctggcgtt cgaatttagc agcagcggtt tctttcatac caattgcagt actaccgcgt
      361 ggcaccagac ccgcggaacc tggtgctgta ccaagaactt tagctccacc tatatatcct
      421 gtaacttttg gatttacaac tggtgttgaa ctacttgcat caaatgaata gtcatttgtt
      481 tgagtgtagt ttgaccagtc tgattttgtt attcttcctt gaatagttat aaattgtcct
      541 tttttaagag tagctgctcc gcttgcaaat ccaaattcaa catatgtatc ataggttgat
      601 gttgggcttg ctgtttcttt aacgaagttt gctgtcactt tgctagtgtt atcaacatag
      661 ctatttccta ataatgcacc agcatggtca caccagaaag tttgtccttg tgtaccatca
      721 cttgtgtaat aatatctaac ttttacgtca tttaaattta aatcactgtc agatgtgtta
      781 gtaattttga ttattggtgt aattgagttt gtttgtgctg atttgttaga gttgtaaaat
      841 tcaactgaca ttatgtatat ctccttctta aagttaaaca aaattatttc tagaggggaa
      901 ttgttatccg ctcacaattc ccctatagtg agtcgtatta atttcgcggg atcgagatcg
      961 atctcgatcc tctacgccgg acgcatcgtg gccggcatca ccggcgccac aggtgcggtt
     1021 gctggcgcct atatcgccga catcaccgat ggggaagatc gggctcgcca cttcgggctc
     1081 atgagcgctt gtttcggcgt gggtatggtg gcaggccccg tggccggggg actgttgggc
     1141 gccatctcct tgcatgcacc attccttgcg gcggcggtgc tcaacggcct caacctacta
     1201 ctgggctgct tcctaatgca ggagtcgcat aagggagagc gtcgagatcc cggacaccat
     1261 cgaatggcgc aaaacctttc gcggtatggc atgatagcgc ccggaagaga gtcaattcag
     1321 ggtggtgaat gtgaaaccag taacgttata cgatgtcgca gagtatgccg gtgtctctta
     1381 tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa cgcgggaaaa
     1441 agtggaagcg gcgatggcgg agctgaatta cattcccaac cgcgtggcac aacaactggc
     1501 gggcaaacag tcgttgctga ttggcgttgc cacctccagt ctggccctgc acgcgccgtc
     1561 gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg tggtggtgtc
     1621 gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc ttctcgcgca
     1681 acgcgtcagt gggctgatca ttaactatcc gctggatgac caggatgcca ttgctgtgga
     1741 agctgcctgc actaatgttc cggcgttatt tcttgatgtc tctgaccaga cacccatcaa
     1801 cagtattatt ttctcccatg aagacggtac gcgactgggc gtggagcatc tggtcgcatt
     1861 gggtcaccag caaatcgcgc tgttagcggg cccattaagt tctgtctcgg cgcgtctgcg
     1921 tctggctggc tggcataaat atctcactcg caatcaaatt cagccgatag cggaacggga
     1981 aggcgactgg agtgccatgt ccggttttca acaaaccatg caaatgctga atgagggcat
     2041 cgttcccact gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa tgcgcgccat
     2101 taccgagtcc gggctgcgcg ttggtgcgga catctcggta gtgggatacg acgataccga
     2161 agacagctca tgttatatcc cgccgttaac caccatcaaa caggattttc gcctgctggg
     2221 gcaaaccagc gtggaccgct tgctgcaact ctctcagggc caggcggtga agggcaatca
     2281 gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata cgcaaaccgc
     2341 ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga
     2401 aagcgggcag tgagcgcaac gcaattaatg taagttagct cactcattag gcaccgggat
     2461 ctcgaccgat gcccttgaga gccttcaacc cagtcagctc cttccggtgg gcgcggggca
     2521 tgactatcgt cgccgcactt atgactgtct tctttatcat gcaactcgta ggacaggtgc
     2581 cggcagcgct ctgggtcatt ttcggcgagg accgctttcg ctggagcgcg acgatgatcg
     2641 gcctgtcgct tgcggtattc ggaatcttgc acgccctcgc tcaagccttc gtcactggtc
     2701 ccgccaccaa acgtttcggc gagaagcagg ccattatcgc cggcatggcg gccccacggg
     2761 tgcgcatgat cgtgctcctg tcgttgagga cccggctagg ctggcggggt tgccttactg
     2821 gttagcagaa tgaatcaccg atacgcgagc gaacgtgaag cgactgctgc tgcaaaacgt
     2881 ctgcgacctg agcaacaaca tgaatggtct tcggtttccg tgtttcgtaa agtctggaaa
     2941 cgcggaagtc agcgccctgc accattatgt tccggatctg catcgcagga tgctgctggc
     3001 taccctgtgg aacacctaca tctgtattaa cgaagcgctg gcattgaccc tgagtgattt
     3061 ttctctggtc ccgccgcatc cataccgcca gttgtttacc ctcacaacgt tccagtaacc
     3121 gggcatgttc atcatcagta acccgtatcg tgagcatcct ctctcgtttc atcggtatca
     3181 ttacccccat gaacagaaat cccccttaca cggaggcatc agtgaccaaa caggaaaaaa
     3241 ccgcccttaa catggcccgc tttatcagaa gccagacatt aacgcttctg gagaaactca
     3301 acgagctgga cgcggatgaa caggcagaca tctgtgaatc gcttcacgac cacgctgatg
     3361 agctttaccg cagctgcctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc
     3421 agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc
     3481 agggcgcgtc agcgggtgtt ggcgggtgtc ggggcgcagc catgacccag tcacgtagcg
     3541 atagcggagt gtatactggc ttaactatgc ggcatcagag cagattgtac tgagagtgca
     3601 ccatatatgc ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg catcaggcgc
     3661 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     3721 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     3781 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     3841 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     3901 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     3961 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     4021 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     4081 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     4141 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     4201 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     4261 cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct gaagccagtt
     4321 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     4381 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     4441 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     4501 gtcatgaaca ataaaactgt ctgcttacat aaacagtaat acaaggggtg ttatgagcca
     4561 tattcaacgg gaaacgtctt gctctaggcc gcgattaaat tccaacatgg atgctgattt
     4621 atatgggtat aaatgggctc gcgataatgt cgggcaatca ggtgcgacaa tctatcgatt
     4681 gtatgggaag cccgatgcgc cagagttgtt tctgaaacat ggcaaaggta gcgttgccaa
     4741 tgatgttaca gatgagatgg tcagactaaa ctggctgacg gaatttatgc ctcttccgac
     4801 catcaagcat tttatccgta ctcctgatga tgcatggtta ctcaccactg cgatccccgg
     4861 gaaaacagca ttccaggtat tagaagaata tcctgattca ggtgaaaata ttgttgatgc
     4921 gctggcagtg ttcctgcgcc ggttgcattc gattcctgtt tgtaattgtc cttttaacag
     4981 cgatcgcgta tttcgtctcg ctcaggcgca atcacgaatg aataacggtt tggttgatgc
     5041 gagtgatttt gatgacgagc gtaatggctg gcctgttgaa caagtctgga aagaaatgca
     5101 taaacttttg ccattctcac cggattcagt cgtcactcat ggtgatttct cacttgataa
     5161 ccttattttt gacgagggga aattaatagg ttgtattgat gttggacgag tcggaatcgc
     5221 agaccgatac caggatcttg ccatcctatg gaactgcctc ggtgagtttt ctccttcatt
     5281 acagaaacgg ctttttcaaa aatatggtat tgataatcct gatatgaata aattgcagtt
     5341 tcatttgatg ctcgatgagt ttttctaaga attaattcat gagcggatac atatttgaat
     5401 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     5461 aaattgtaaa cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
     5521 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
     5581 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
     5641 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct
     5701 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc
     5761 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag
     5821 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca
     5881 cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgcca
//

      

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