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基本信息

質粒圖譜和多克隆位點信息 

注意：

客戶在我公司選擇構建載體服務項目時，請自行提供母源載體。

如要使用本公司保存的母源載體時，本公司不能保證所有母源載體完全符合客戶預期，也不承擔由此產生的任何責任。

載體描述

      pET-30a(+)載體攜帶有一個N端的His標簽/Thrombin酶切位點/ S標簽/ EK蛋白酶切標簽以及一個可選擇的C端His標簽。 

      載體的多克隆位點見上面的圖譜。需要注意的是圖譜中的核苷酸編碼順序是以pBR322載體的復制方向為正方向，所以T7表達區域在圖譜中是處于反向的位 置。T7 RNA聚合酶啟動的克隆和表達區域在質粒圖譜中也被標注了出來。質粒的F1復制子是被定向的，所以在T7噬菌體聚合酶的作用下，包含有蛋白編碼序列的病毒 粒子能夠產生，并啟動蛋白表達，同時蛋白表達將被T7終止子序列(Cat. No. 69337-3)的作用下終止蛋白翻譯。

載體序列

LOCUS       Exported                5422 bp ds-DNA     circular SYN 14-MAR-2019

DEFINITION  synthetic circular DNA

ACCESSION   .

VERSION     .

KEYWORDS    .

SOURCE      synthetic DNA construct

  ORGANISM  synthetic DNA construct

REFERENCE   1  (bases 1 to 5422)

  AUTHORS   .

  TITLE     Direct Submission

  JOURNAL   Exported Thursday, Mar 14, 2019 from SnapGene Viewer 4.3.4

FEATURES             Location/Qualifiers

     source          1..5422

                     /organism="synthetic DNA construct"

                     /mol_type="other DNA"

     rep_origin      12..467

                     /direction=RIGHT

                     /label=f1 ori

                     /note="f1 bacteriophage origin of replication; arrow

                     indicates direction of (+) strand synthesis"

     CDS             complement(560..1375)

                     /codon_start=1

                     /gene="aph(3')-Ia"

                     /product="aminoglycoside phosphotransferase"

                     /label=KanR

                     /note="confers resistance to kanamycin in bacteria or G418

                     (Geneticin(R)) in eukaryotes"

                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG

                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK

                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA

                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI

                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"

     rep_origin      1497..2085

                     /direction=RIGHT

                     /label=ori

                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of

                     replication"

     CDS             complement(2515..2706)

                     /codon_start=1

                     /gene="rop"

                     /product="Rop protein, which maintains plasmids at low copy

                     number"

                     /label=rop

                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA

                     DELYRSCLARFGDDGENL"

     CDS             complement(3515..4597)

                     /codon_start=1

                     /gene="lacI"

                     /product="lac repressor"

                     /label=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"

     promoter        complement(4598..4675)

                     /gene="lacI"

                     /label=lacI promoter

     primer_bind     4988..5007

                     /label=T7 primer

     promoter        4988..5006

                     /label=T7 promoter

                     /note="promoter for bacteriophage T7 RNA polymerase"

     protein_bind    5007..5031

                     /label=lac operator

                     /bound_moiety="lac repressor encoded by 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)."

     RBS             5046..5068

                     /note="efficient ribosome binding site from bacteriophage

                     T7 gene 10 (Olins and Rangwala, 1989)"

     CDS             5079..5096

                     /codon_start=1

                     /product="6xHis affinity tag"

                     /label=6xHis

                     /translation="HHHHHH"

     CDS             5106..5123

                     /codon_start=1

                     /product="thrombin recognition and cleavage site"

                     /label=thrombin site

                     /translation="LVPRGS"

     CDS             5130..5174

                     /codon_start=1

                     /product="affinity and epitope tag derived from pancreatic

                     ribonuclease A"

                     /label=S-Tag

                     /translation="KETAAAKFERQHMDS"

     CDS             5190..5204

                     /codon_start=1

                     /product="enterokinase recognition and cleavage site"

                     /label=enterokinase site

                     /translation="DDDDK"

     misc_feature    5206..5265

                     /label=MCS

     CDS             5266..5283

                     /codon_start=1

                     /product="6xHis affinity tag"

                     /label=6xHis

                     /translation="HHHHHH"

     primer_bind     complement(5336..5354)

                     /label=T7 ter primer

     terminator      5350..5397

                     /label=T7 terminator

                     /note="transcription terminator for bacteriophage T7 RNA

                     polymerase"

ORIGIN

        1 tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg

       61 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc

      121 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg

      181 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc

      241 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt

      301 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc

      361 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta

      421 acaaaaattt aacgcgaatt ttaacaaaat attaacgttt acaatttcag gtggcacttt

      481 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta

      541 tccgctcatg aattaattct tagaaaaact catcgagcat caaatgaaac tgcaatttat

      601 tcatatcagg attatcaata ccatattttt gaaaaagccg tttctgtaat gaaggagaaa

      661 actcaccgag gcagttccat aggatggcaa gatcctggta tcggtctgcg attccgactc

      721 gtccaacatc aatacaacct attaatttcc cctcgtcaaa aataaggtta tcaagtgaga

      781 aatcaccatg agtgacgact gaatccggtg agaatggcaa aagtttatgc atttctttcc

      841 agacttgttc aacaggccag ccattacgct cgtcatcaaa atcactcgca tcaaccaaac

      901 cgttattcat tcgtgattgc gcctgagcga gacgaaatac gcgatcgctg ttaaaaggac

      961 aattacaaac aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca tcaacaatat

     1021 tttcacctga atcaggatat tcttctaata cctggaatgc tgttttcccg gggatcgcag

     1081 tggtgagtaa ccatgcatca tcaggagtac ggataaaatg cttgatggtc ggaagaggca

     1141 taaattccgt cagccagttt agtctgacca tctcatctgt aacatcattg gcaacgctac

     1201 ctttgccatg tttcagaaac aactctggcg catcgggctt cccatacaat cgatagattg

     1261 tcgcacctga ttgcccgaca ttatcgcgag cccatttata cccatataaa tcagcatcca

     1321 tgttggaatt taatcgcggc ctagagcaag acgtttcccg ttgaatatgg ctcataacac

     1381 cccttgtatt actgtttatg taagcagaca gttttattgt tcatgaccaa aatcccttaa

     1441 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga

     1501 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg

     1561 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc

     1621 agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag

     1681 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc

     1741 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg

     1801 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac

     1861 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga

     1921 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt

     1981 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag

     2041 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg

     2101 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta

     2161 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc

     2221 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg

     2281 tattttctcc ttacgcatct gtgcggtatt tcacaccgca tatatggtgc actctcagta

     2341 caatctgctc tgatgccgca tagttaagcc agtatacact ccgctatcgc tacgtgactg

     2401 ggtcatggct gcgccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct

     2461 gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag

     2521 gttttcaccg tcatcaccga aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc

     2581 gtgaagcgat tcacagatgt ctgcctgttc atccgcgtcc agctcgttga gtttctccag

     2641 aagcgttaat gtctggcttc tgataaagcg ggccatgtta agggcggttt tttcctgttt

     2701 ggtcactgat gcctccgtgt aagggggatt tctgttcatg ggggtaatga taccgatgaa

     2761 acgagagagg atgctcacga tacgggttac tgatgatgaa catgcccggt tactggaacg

     2821 ttgtgagggt aaacaactgg cggtatggat gcggcgggac cagagaaaaa tcactcaggg

     2881 tcaatgccag cgcttcgtta atacagatgt aggtgttcca cagggtagcc agcagcatcc

     2941 tgcgatgcag atccggaaca taatggtgca gggcgctgac ttccgcgttt ccagacttta

     3001 cgaaacacgg aaaccgaaga ccattcatgt tgttgctcag gtcgcagacg ttttgcagca

     3061 gcagtcgctt cacgttcgct cgcgtatcgg tgattcattc tgctaaccag taaggcaacc

     3121 ccgccagcct agccgggtcc tcaacgacag gagcacgatc atgcgcaccc gtggggccgc

     3181 catgccggcg ataatggcct gcttctcgcc gaaacgtttg gtggcgggac cagtgacgaa

     3241 ggcttgagcg agggcgtgca agattccgaa taccgcaagc gacaggccga tcatcgtcgc

     3301 gctccagcga aagcggtcct cgccgaaaat gacccagagc gctgccggca cctgtcctac

     3361 gagttgcatg ataaagaaga cagtcataag tgcggcgacg atagtcatgc cccgcgccca

     3421 ccggaaggag ctgactgggt tgaaggctct caagggcatc ggtcgagatc ccggtgccta

     3481 atgagtgagc taacttacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa

     3541 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat

     3601 tgggcgccag ggtggttttt cttttcacca gtgagacggg caacagctga ttgcccttca

     3661 ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct ggtttgcccc agcaggcgaa

     3721 aatcctgttt gatggtggtt aacggcggga tataacatga gctgtcttcg gtatcgtcgt

     3781 atcccactac cgagatgtcc gcaccaacgc gcagcccgga ctcggtaatg gcgcgcattg

     3841 cgcccagcgc catctgatcg ttggcaacca gcatcgcagt gggaacgatg ccctcattca

     3901 gcatttgcat ggtttgttga aaaccggaca tggcactcca gtcgccttcc cgttccgcta

     3961 tcggctgaat ttgattgcga gtgagatatt tatgccagcc agccagacgc agacgcgccg

     4021 agacagaact taatgggccc gctaacagcg cgatttgctg gtgacccaat gcgaccagat

     4081 gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat aatactgttg atgggtgtct

     4141 ggtcagagac atcaagaaat aacgccggaa cattagtgca ggcagcttcc acagcaatgg

     4201 catcctggtc atccagcgga tagttaatga tcagcccact gacgcgttgc gcgagaagat

     4261 tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc taccatcgac accaccacgc

     4321 tggcacccag ttgatcggcg cgagatttaa tcgccgcgac aatttgcgac ggcgcgtgca

     4381 gggccagact ggaggtggca acgccaatca gcaacgactg tttgcccgcc agttgttgtg

     4441 ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc ttccactttt tcccgcgttt

     4501 tcgcagaaac gtggctggcc tggttcacca cgcgggaaac ggtctgataa gagacaccgg

     4561 catactctgc gacatcgtat aacgttactg gtttcacatt caccaccctg aattgactct

     4621 cttccgggcg ctatcatgcc ataccgcgaa aggttttgcg ccattcgatg gtgtccggga

     4681 tctcgacgct ctcccttatg cgactcctgc attaggaagc agcccagtag taggttgagg

     4741 ccgttgagca ccgccgccgc aaggaatggt gcatgcaagg agatggcgcc caacagtccc

     4801 ccggccacgg ggcctgccac catacccacg ccgaaacaag cgctcatgag cccgaagtgg

     4861 cgagcccgat cttccccatc ggtgatgtcg gcgatatagg cgccagcaac cgcacctgtg

     4921 gcgccggtga tgccggccac gatgcgtccg gcgtagagga tcgagatcga tctcgatccc

     4981 gcgaaattaa tacgactcac tataggggaa ttgtgagcgg ataacaattc ccctctagaa

     5041 ataattttgt ttaactttaa gaaggagata tacatatgca ccatcatcat catcattctt

     5101 ctggtctggt gccacgcggt tctggtatga aagaaaccgc tgctgctaaa ttcgaacgcc

     5161 agcacatgga cagcccagat ctgggtaccg acgacgacga caaggccatg gctgatatcg

     5221 gatccgaatt cgagctccgt cgacaagctt gcggccgcac tcgagcacca ccaccaccac

     5281 cactgagatc cggctgctaa caaagcccga aaggaagctg agttggctgc tgccaccgct

     5341 gagcaataac tagcataacc ccttggggcc tctaaacggg tcttgagggg ttttttgctg

     5401 aaaggaggaa ctatatccgg at

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