LOCUS       sequence-67097-e        6553 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  FX cloning E.coli expression vector with araBAD promoter and 
            C-terminal Avi-tag, 3C protease cleavage site, GFP and 10x His tags.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6553)
  AUTHORS   Geertsma ER, Dutzler R
  TITLE     A versatile and efficient high-throughput cloning tool for 
            structural biology.
  JOURNAL   Biochemistry. 2011 Apr 19;50(15):3272-8. Epub 2011 Mar 25.
  PUBMED    21410291
REFERENCE   2  (bases 1 to 6553)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Biochemistry."; date: "2011-04-19"; volume: "50(15)"; pages: 
            "3272-8. Epub 2011 Mar 25"
COMMENT     Sequence Label: pBXCA3GH
FEATURES             Location/Qualifiers
     source          1..6553
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..285
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     208..227
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             complement(370..675)
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     complement(472..491)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(1017..1676)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     1591..1610
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(join(1730..1736,1737..1754,1755..1760))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(1785..1909)
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(1928..1948)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(1952..1972)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1999..2043
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             2044..2067
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2074..2784
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Nager et al., 2011)"
                     /label=superfolder GFP
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKAN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     primer_bind     complement(2101..2129)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     2724..2744
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     CDS             2791..2820
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             2836..2865
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2881..2898
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2954..2971)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      3124..3210
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3302..3329
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        3348..3439
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3440..3508,3509..4300)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3658..3677)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4471..5059
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4960..4979
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5213..5230
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5245..5385
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5311..5330)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5471..5493
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5649..6527)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
ORIGIN
        1 aagaaaccaa ttgtccatat tgcatcagac attgccgtca ctgcgtcttt tactggcact
       61 tctcgctaac caaaccggta accccgctta ttaaaagcat tctgtaacaa agcgggacca
      121 aagccatgac aaaaacgcgt aacaaaagtg tctataatca cggcagaaaa gtccacattg
      181 attatttgca cggcgtcaca ctttgctatg ccatagcatt tttatccata agattagcgg
      241 atcctacctg acgcttttta tcgcaactct ctactgtttc tccatacccg ttttttgggc
      301 taacaggagg aattaaccat gagttgaaga gcgacctgca gactggctgt gtataaggga
      361 gcctgacatt tatattcccc agaacatcag gttaatggcg tttttgatgt cattttcgcg
      421 gtggctgaga tcagccactt cttccccgat aacggagacc ggcacactgg ccatatcggt
      481 ggtcatcatg cgccagcttt catccccgat atgcaccacc gggtaaagtt cacgggagac
      541 tttatctgac agcagacgtg cactggccag ggggatcacc atccgtcgcc cgggcgtgtc
      601 aataatatca ctctgtacat ccacaaacag acgataacgg ctctctcttt tataggtgta
      661 aaccttaaac tgcatttcac cagcccctgt tctcgtcagc aaaagagccg ttcatttcaa
      721 taaaccgggc gacctcagcc atcccttcct gattttccgc tttccagcgt tcggcacgca
      781 gacgacgggc ttcattctgc atggttgtgc ttaccagacc ggagatattg acatcatata
      841 tgccttgagc aactgatagc tgtcgctgtc aactgtcact gtaatacgct gcttcatagc
      901 atacctcttt ttgacatact tcgggtatac atatcagtat atattcttat accgcaaaaa
      961 tcagcgcgca aatacgcata ctgttatctg gcttttagta agccggatcc acgcgtttac
     1021 gccccgccct gccactcatc gcagtactgt tgtaattcat taagcattct gccgacatgg
     1081 aagccatcac aaacggcatg atgaacctga atcgccagcg gcatcagcac cttgtcgcct
     1141 tgcgtataat atttgcccat ggtgaaaacg ggggcgaaga agttgtccat attggccacg
     1201 tttaaatcaa aactggtgaa actcacccag ggattggctg agacgaaaaa catattctca
     1261 ataaaccctt tagggaaata ggccaggttt tcaccgtaac acgccacatc ttgcgaatat
     1321 atgtgtagaa actgccggaa atcgtcgtgg tattcactcc agagcgatga aaacgtttca
     1381 gtttgctcat ggaaaacggt gtaacaaggg tgaacactat cccatatcac cagctcaccg
     1441 tctttcattg ccatacggaa ttccggatga gcattcatca ggcgggcaag aatgtgaata
     1501 aaggccggat aaaacttgtg cttatttttc tttacggtct ttaaaaaggc cgtaatatcc
     1561 agctgaacgg tctggttata ggtacattga gcaactgact gaaatgcctc aaaatgttct
     1621 ttacgatgcc attgggatat atcaacggtg gtatatccag tgattttttt ctccatttta
     1681 gcttccttag ctcctgaaaa tctcgacgga tcctaactca aaatccacac attatacgag
     1741 ccggaagcat aaagtgtaaa gcctggggtg cctaatgcgg ccgccatagt gactggatat
     1801 gttgtgtttt acagtattat gtagtctgtt ttttatgcaa aatctaattt aatatattga
     1861 tatttatatc attttacgtt tctcgttcag cttttttgta caaacttgtg atatcgaatt
     1921 cgctagctac ctttctcttc ttttttggat ctacctttct cttctttttt ggggatccac
     1981 tagttttgct cttctgcagg tctgaacgat atcttcgaag ctcagaaaat cgaatggcac
     2041 gaattagaag ttttgtttca aggtccacaa ttcagcaaag gagaagaact tttcactgga
     2101 gttgtcccaa ttcttgttga attagatggt gatgttaatg ggcacaaatt ttctgtcagt
     2161 ggagagggtg aaggtgatgc tacatacgga aaactcaccc ttaaatttat ttgcactact
     2221 ggaaaactac ctgttccttg gccaacactt gtcactactc tgacctatgg tgttcaatgc
     2281 ttttcccgtt atccggatca catgaaacgg catgactttt tcaagagtgc catgcccgaa
     2341 ggttatgtac aggaacgcac tatatctttc aaagatgacg ggaactacaa gacgcgtgct
     2401 gaagtcaagt ttgaaggtga tacccttgtt aatcgtatcg agttaaaggg tattgatttt
     2461 aaagaagatg gaaacattct cggacacaaa ctagagtaca actataactc acacaatgta
     2521 tacatcacgg cagacaaaca aaagaatgga atcaaagcta acttcaaaat tcgccacaac
     2581 attgaagatg gttccgttca actagcagac cattatcaac aaaatactcc aattggcgat
     2641 ggccctgtcc ttttaccaga caaccattac ctgtcgacac aatctgccct ttcgaaagat
     2701 cccaacgaaa agcgtgacca catggtcctt cttgagtttg taactgctgc tgggattaca
     2761 catggcatgg atgagctcta caaaactagt catcatcatc accatcatca tcaccatcat
     2821 taagtcgacc atctagaaca aaaactcatc tcagaagagg atctgaatag cgccgtcgac
     2881 catcatcatc atcatcattg agtttaaacg gtctccagct tggctgtttt ggcggatgag
     2941 agaagatttt cagcctgata cagattaaat cagaacgcag aagcggtctg ataaaacaga
     3001 atttgcctgg cggcagtagc gcggtggtcc cacctgaccc catgccgaac tcagaagtga
     3061 aacgccgtag cgccgatggt agtgtggggt ctccccatgc gagagtaggg aactgccagg
     3121 catcaaataa aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg
     3181 tcggtgaacg ctctcctgag taggacaaat ccgccgggag cggatttgaa cgttgcgaag
     3241 caacggcccg gagggtggcg ggcaggacgc ccgccataaa ctgccaggca tcaaattaag
     3301 cagaaggcca tcctgacgga tggccttttt gcgtttctac aaactctttt gtttattttt
     3361 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     3421 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt
     3481 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     3541 tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat
     3601 ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct
     3661 atgtggcgcg gtattatccc gtgttgacgc cgggcaagag caactcggtc gccgcataca
     3721 ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg
     3781 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     3841 cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     3901 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     3961 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg
     4021 cgaactactt actctagctt cccggcaaca attaatagac tggatggagg cggataaagt
     4081 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     4141 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     4201 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     4261 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc
     4321 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     4381 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     4441 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     4501 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     4561 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct
     4621 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     4681 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     4741 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     4801 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     4861 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     4921 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     4981 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     5041 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     5101 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
     5161 taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc
     5221 agtgagcgag gaagcggaag tgcccctgat gcggtatttt ctccttacgc atctgtgcgg
     5281 tatttcacac cgcatatggt gcactctcag tacaatctgc tctgatgccg catagttaag
     5341 ccagtataca ctccgctatc gctacgtgac tgggtcatgg ctgcgccccg acacccgcca
     5401 acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct
     5461 gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg
     5521 aggcagcaga tcaattcgcg cgcgaaggcg aagcggcatg cataatgtgc ctgtcaaatg
     5581 gacgaagcag ggattctgca aaccctatgc tactccgtca agccgtcaat tgtctgattc
     5641 gttaccaatt atgacaactt gacggctaca tcattcactt tttcttcaca accggcacgg
     5701 aactcgctcg ggctggcccc ggtgcatttt ttaaataccc gcgagaaata gagttgatcg
     5761 tcaaaaccaa cattgcgacc gacggtggcg ataggcatcc gggtggtgct caaaagcagc
     5821 ttcgcctggc tgatacgttg gtcctcgcgc cagcttaaga cgctaatccc taactgctgg
     5881 cggaaaagat gtgacagacg cgacggcgac aagcaaacat gctgtgcgac gctggcgata
     5941 tcaaaattgc tgtctgccag gtgatcgctg atgtactgac aagcctcgcg tacccgatta
     6001 tccatcggtg gatggagcga ctcgttaatc gcttccatgc gccgcagtaa caattgctca
     6061 agcagattta tcgccagcag ctccgaatag cgcccttccc cttgcccggc gttaatgatt
     6121 tgcccaaaca ggtcgctgaa atgcggctgg tgcgcttcat ccgggcgaaa gaaccccgta
     6181 ttggcaaata ttgacggcca gttaagccat tcatgccagt aggcgcgcgg acgaaagtaa
     6241 acccactggt gataccattc gcgagcctcc ggatgacgac cgtagtgatg aatctctcct
     6301 ggcgggaaca gcaaaatatc acccggtcgg caaacaaatt ctcgtccctg atttttcacc
     6361 accccctgac cgcgaatggt gagattgaga atataacctt tcattcccag cggtcggtcg
     6421 ataaaaaaat cgagataacc gttggcctca atcggcgtta aacccgccac cagatgggca
     6481 ttaaacgagt atcccggcag caggggatca ttttgcgctt cagccatact tttcatactc
     6541 ccgccattca gag
//
