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載體描述

pCDNA3.1-his-EGFP  載體在pCDNA3.1-EGFP的基礎上添加了10xHis標簽，以便于目的蛋白純化。

載體序列

LOCUS       Exported                6160 bp ds-DNA     circular SYN 27-AUG-2019

DEFINITION  synthetic circular DNA

ACCESSION   .

VERSION     .

KEYWORDS    pCDNA3.1-his-EGFP

SOURCE      synthetic DNA construct

  ORGANISM  synthetic DNA construct

REFERENCE   1  (bases 1 to 6160)

  AUTHORS   .

  TITLE     Direct Submission

  JOURNAL   Exported Aug 27, 2019 from SnapGene Viewer 4.2.6

            http://www.snapgene.com

FEATURES             Location/Qualifiers

     source          1..6160

                     /organism="synthetic DNA construct"

                     /mol_type="other DNA"

     enhancer        235..614

                     /label=CMV enhancer

                     /note="human cytomegalovirus immediate early enhancer"

     promoter        615..818

                     /label=CMV promoter

                     /note="human cytomegalovirus (CMV) immediate early

                     promoter"

     promoter        863..881

                     /label=T7 promoter

                     /note="promoter for bacteriophage T7 RNA polymerase"

     misc_feature    895..957

                     /label=MCS

     CDS             967..996

                     /codon_start=1

                     /product="10xHis affinity tag"

                     /label=10xHis

                     /translation="HHHHHHHHHH"

     CDS             1003..1722

                     /codon_start=1

                     /product="enhanced GFP"

                     /label=EGFP

                     /note="mammalian codon-optimized"

                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL

                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD

                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK

                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL

                     EFVTAAGITLGMDELYK"

     polyA_signal    1760..1984

                     /label=bGH poly(A) signal

                     /note="bovine growth hormone polyadenylation signal"

     rep_origin      2030..2458

                     /direction=RIGHT

                     /label=f1 ori

                     /note="f1 bacteriophage origin of replication; arrow

                     indicates direction of (+) strand synthesis"

     promoter        2472..2801

                     /label=SV40 promoter

                     /note="SV40 enhancer and early promoter"

     rep_origin      2652..2787

                     /label=SV40 ori

                     /note="SV40 origin of replication"

     CDS             2868..3662

                     /codon_start=1

                     /gene="aph(3')-II (or nptII)"

                     /product="aminoglycoside phosphotransferase from Tn5"

                     /label=NeoR/KanR

                     /note="confers resistance to neomycin, kanamycin, and G418

                     (Geneticin(R))"

                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP

                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS

                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ

                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA

                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"

     polyA_signal    3836..3957

                     /label=SV40 poly(A) signal

                     /note="SV40 polyadenylation signal"

     primer_bind     complement(4006..4022)

                     /label=M13 rev

                     /note="common sequencing primer, one of multiple similar

                     variants"

     protein_bind    4030..4046

                     /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)."

     promoter        complement(4054..4084)

                     /label=lac promoter

                     /note="promoter for the E. coli lac operon"

     protein_bind    4099..4120

                     /label=CAP binding site

                     /bound_moiety="E. coli catabolite activator protein"

                     /note="CAP binding activates transcription in the presence

                     of cAMP."

     rep_origin      complement(4408..4993)

                     /direction=LEFT

                     /label=ori

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

                     replication"

     CDS             complement(5164..6024)

                     /codon_start=1

                     /gene="bla"

                     /product="beta-lactamase"

                     /label=AmpR

                     /note="confers resistance to ampicillin, carbenicillin, and

                     related antibiotics"

                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI

                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS

                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW

                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA

                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS

                     LIKHW"

     promoter        complement(6025..6129)

                     /gene="bla"

                     /label=AmpR promoter

ORIGIN

        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg

       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg

      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc

      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt

      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata

      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc

      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc

      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt

      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt

      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca

      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg

      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc

      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg

      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca

      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc

      901 gtctaaactt aagcttggta ccgagctcgg atccactagt ccagtgtggt ggaattcgcc

      961 accatgcatc atcatcatca tcatcatcat catcatctcg agatggtgag caagggcgag

     1021 gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac

     1081 aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag

     1141 ttcatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc

     1201 tacggcgtgc agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag

     1261 tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac

     1321 tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg

     1381 aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac

     1441 aacagccaca acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc

     1501 aagatccgcc acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac

     1561 acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc

     1621 gccctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc

     1681 gccgccggga tcactctcgg catggacgag ctgtacaagt aatctagagg gcccgtttaa

     1741 acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc

     1801 cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag

     1861 gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag

     1921 gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct

     1981 atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt

     2041 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc

     2101 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc

     2161 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg

     2221 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga

     2281 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc

     2341 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg

     2401 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa

     2461 ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa

     2521 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc

     2581 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc

     2641 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct

     2701 gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga

     2761 agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta

     2821 tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg attgaacaag

     2881 atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg

     2941 cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc

     3001 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag

     3061 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca

     3121 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat

     3181 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata

     3241 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac

     3301 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc

     3361 tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg

     3421 tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg

     3481 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta

     3541 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg

     3601 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct

     3661 gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga

     3721 tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc

     3781 cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt

     3841 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa

     3901 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca

     3961 tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat agctgtttcc

     4021 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg

     4081 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc

     4141 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg

     4201 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc

     4261 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac

     4321 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa

     4381 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca

     4441 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc

     4501 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata

     4561 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta

     4621 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca

     4681 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga

     4741 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg

     4801 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg

     4861 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg

     4921 caaacaaacc accgctggta gcggtttttt tgtttgcaag cagcagatta cgcgcagaaa

     4981 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga

     5041 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct

     5101 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga

     5161 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc

     5221 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg

     5281 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat

     5341 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat

     5401 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg

     5461 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc

     5521 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa

     5581 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc

     5641 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt

     5701 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag

     5761 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt

     5821 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag

     5881 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac

     5941 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc

     6001 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca

     6061 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg

     6121 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc

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