LOCUS       Exported                5824 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  FX cloning E.coli expression vector with araBAD promoter, N-terminal
            10x His tag, 3C protease cleavage site and C-terminal Avi-tag.
ACCESSION   .
VERSION     .
KEYWORDS    pBXNH3CA
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5824)
  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 5824)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5824
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     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             322..351
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             352..375
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             complement(424..729)
                     /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(526..545)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(1071..1730)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     1645..1664
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(1784..1814)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(1839..1963)
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(1982..2002)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(2006..2026)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2053..2097
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             2107..2136
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2152..2169
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2225..2242)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      2395..2481
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2573..2600
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        2619..2710
                     /gene="bla"
                     /label=AmpR promoter
     CDS             2711..3571
                     /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(2929..2948)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3742..4330
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4231..4250
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4484..4501
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4516..4656
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4582..4601)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4742..4764
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4920..5798)
                     /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 tactggctct
       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 gcaccatcat catcaccatc atcatcatca tttagaagtt
      361 ttgtttcaag gtccaagtag aagagcgagc tgcagactgg ctgtgtataa gggagcctga
      421 catttatatt ccccagaaca tcaggttaat ggcgtttttg atgtcatttt cgcggtggct
      481 gagatcagcc acttcttccc cgataacgga gaccggcaca ctggccatat cggtggtcat
      541 catgcgccag ctttcatccc cgatatgcac caccgggtaa agttcacggg agactttatc
      601 tgacagcaga cgtgcactgg ccagggggat caccatccgt cgcccgggcg tgtcaataat
      661 atcactctgt acatccacaa acagacgata acggctctct cttttatagg tgtaaacctt
      721 aaactgcatt tcaccagccc ctgttctcgt cagcaaaaga gccgttcatt tcaataaacc
      781 gggcgacctc agccatccct tcctgatttt ccgctttcca gcgttcggca cgcagacgac
      841 gggcttcatt ctgcatggtt gtgcttacca gaccggagat attgacatca tatatgcctt
      901 gagcaactga tagctgtcgc tgtcaactgt cactgtaata cgctgcttca tagcatacct
      961 ctttttgaca tacttcgggt atacatatca gtatatattc ttataccgca aaaatcagcg
     1021 cgcaaatacg catactgtta tctggctttt agtaagccgg atccacgcgt ttacgccccg
     1081 ccctgccact catcgcagta ctgttgtaat tcattaagca ttctgccgac atggaagcca
     1141 tcacaaacgg catgatgaac ctgaatcgcc agcggcatca gcaccttgtc gccttgcgta
     1201 taatatttgc ccatggtgaa aacgggggcg aagaagttgt ccatattggc cacgtttaaa
     1261 tcaaaactgg tgaaactcac ccagggattg gctgagacga aaaacatatt ctcaataaac
     1321 cctttaggga aataggccag gttttcaccg taacacgcca catcttgcga atatatgtgt
     1381 agaaactgcc ggaaatcgtc gtggtattca ctccagagcg atgaaaacgt ttcagtttgc
     1441 tcatggaaaa cggtgtaaca agggtgaaca ctatcccata tcaccagctc accgtctttc
     1501 attgccatac ggaattccgg atgagcattc atcaggcggg caagaatgtg aataaaggcc
     1561 ggataaaact tgtgcttatt tttctttacg gtctttaaaa aggccgtaat atccagctga
     1621 acggtctggt tataggtaca ttgagcaact gactgaaatg cctcaaaatg ttctttacga
     1681 tgccattggg atatatcaac ggtggtatat ccagtgattt ttttctccat tttagcttcc
     1741 ttagctcctg aaaatctcga cggatcctaa ctcaaaatcc acacattata cgagccggaa
     1801 gcataaagtg taaagcctgg ggtgcctaat gcggccgcca tagtgactgg atatgttgtg
     1861 ttttacagta ttatgtagtc tgttttttat gcaaaatcta atttaatata ttgatattta
     1921 tatcatttta cgtttctcgt tcagcttttt tgtacaaact tgtgatatcg aattcgctag
     1981 ctacctttct cttctttttt ggatctacct ttctcttctt ttttggggat ccactagttt
     2041 tgctcttctg caggtctgaa cgatatcttc gaagctcaga aaatcgaatg gcacgaataa
     2101 taactagaac aaaaactcat ctcagaagag gatctgaata gcgccgtcga ccatcatcat
     2161 catcatcatt gagtttaaac ggtctccagc ttggctgttt tggcggatga gagaagattt
     2221 tcagcctgat acagattaaa tcagaacgca gaagcggtct gataaaacag aatttgcctg
     2281 gcggcagtag cgcggtggtc ccacctgacc ccatgccgaa ctcagaagtg aaacgccgta
     2341 gcgccgatgg tagtgtgggg tctccccatg cgagagtagg gaactgccag gcatcaaata
     2401 aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta tctgttgttt gtcggtgaac
     2461 gctctcctga gtaggacaaa tccgccggga gcggatttga acgttgcgaa gcaacggccc
     2521 ggagggtggc gggcaggacg cccgccataa actgccaggc atcaaattaa gcagaaggcc
     2581 atcctgacgg atggcctttt tgcgtttcta caaactcttt tgtttatttt tctaaataca
     2641 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     2701 aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt
     2761 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca
     2821 gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag
     2881 ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc
     2941 ggtattatcc cgtgttgacg ccgggcaaga gcaactcggt cgccgcatac actattctca
     3001 gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt
     3061 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct
     3121 gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt
     3181 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga
     3241 caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact
     3301 tactctagct tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc
     3361 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga
     3421 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt
     3481 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga
     3541 gataggtgcc tcactgatta agcattggta actgtcagac caagtttact catatatact
     3601 ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga
     3661 taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     3721 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     3781 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     3841 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc ttctagtgta
     3901 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     3961 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     4021 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     4081 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     4141 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     4201 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     4261 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     4321 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     4381 tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt
     4441 tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga
     4501 ggaagcggaa gtgcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca
     4561 ccgcatatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatac
     4621 actccgctat cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc aacacccgct
     4681 gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc
     4741 tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gaggcagcag
     4801 atcaattcgc gcgcgaaggc gaagcggcat gcataatgtg cctgtcaaat ggacgaagca
     4861 gggattctgc aaaccctatg ctactccgtc aagccgtcaa ttgtctgatt cgttaccaat
     4921 tatgacaact tgacggctac atcattcact ttttcttcac aaccggcacg gaactcgctc
     4981 gggctggccc cggtgcattt tttaaatacc cgcgagaaat agagttgatc gtcaaaacca
     5041 acattgcgac cgacggtggc gataggcatc cgggtggtgc tcaaaagcag cttcgcctgg
     5101 ctgatacgtt ggtcctcgcg ccagcttaag acgctaatcc ctaactgctg gcggaaaaga
     5161 tgtgacagac gcgacggcga caagcaaaca tgctgtgcga cgctggcgat atcaaaattg
     5221 ctgtctgcca ggtgatcgct gatgtactga caagcctcgc gtacccgatt atccatcggt
     5281 ggatggagcg actcgttaat cgcttccatg cgccgcagta acaattgctc aagcagattt
     5341 atcgccagca gctccgaata gcgcccttcc ccttgcccgg cgttaatgat ttgcccaaac
     5401 aggtcgctga aatgcggctg gtgcgcttca tccgggcgaa agaaccccgt attggcaaat
     5461 attgacggcc agttaagcca ttcatgccag taggcgcgcg gacgaaagta aacccactgg
     5521 tgataccatt cgcgagcctc cggatgacga ccgtagtgat gaatctctcc tggcgggaac
     5581 agcaaaatat cacccggtcg gcaaacaaat tctcgtccct gatttttcac caccccctga
     5641 ccgcgaatgg tgagattgag aatataacct ttcattccca gcggtcggtc gataaaaaaa
     5701 tcgagataac cgttggcctc aatcggcgtt aaacccgcca ccagatgggc attaaacgag
     5761 tatcccggca gcaggggatc attttgcgct tcagccatac ttttcatact cccgccattc
     5821 agag
//
