LOCUS       Exported                5848 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  E.coli expression vector for FX cloning system, N-terminal pelB 
            signal sequence followed by 10xHisTag and 3C cleavage site.
ACCESSION   .
VERSION     .
KEYWORDS    pBXNPH3
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5848)
  AUTHORS   Zimmermann I, Egloff P, Hutter CA, Arnold FM, Stohler P, Bocquet N, 
            Hug MN, Huber S, Siegrist M, Hetemann L, Gera J, Gmur S, Spies P, 
            Gygax D, Geertsma ER, Dawson RJ, Seeger MA
  TITLE     Synthetic single domain antibodies for the conformational trapping 
            of membrane proteins.
  JOURNAL   Elife. 2018 May 24;7. pii: 34317. doi: 10.7554/eLife.34317.
  PUBMED    29792401
REFERENCE   2  (bases 1 to 5848)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5848
                     /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"
     sig_peptide     319..384
                     /gene="pelB (fragment)"
                     /product="leader peptide for secretion"
                     /label=pelB signal sequence
     CDS             391..420
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             421..444
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             complement(493..798)
                     /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(595..614)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(1140..1799)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     1714..1733
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(1853..1883)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(1908..2032)
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(2051..2071)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(2075..2095)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2131..2160
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2176..2193
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2249..2266)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      2419..2505
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2597..2624
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        2643..2734
                     /gene="bla"
                     /label=AmpR promoter
     CDS             2735..3595
                     /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(2953..2972)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3766..4354
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4255..4274
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4508..4525
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4540..4680
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4606..4625)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4766..4788
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4944..5822)
                     /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 gaaatacctg ctgccgaccg ctgctgccgg actgctgctc
      361 ctcgctgccc agcctgcaat ggccgcagat caccatcatc atcaccatca tcatcatcat
      421 ttagaagttt tgtttcaagg tccaagtaga agagcgagct gcagactggc tgtgtataag
      481 ggagcctgac atttatattc cccagaacat caggttaatg gcgtttttga tgtcattttc
      541 gcggtggctg agatcagcca cttcttcccc gataacggag accggcacac tggccatatc
      601 ggtggtcatc atgcgccagc tttcatcccc gatatgcacc accgggtaaa gttcacggga
      661 gactttatct gacagcagac gtgcactggc cagggggatc accatccgtc gcccgggcgt
      721 gtcaataata tcactctgta catccacaaa cagacgataa cggctctctc ttttataggt
      781 gtaaacctta aactgcattt caccagcccc tgttctcgtc agcaaaagag ccgttcattt
      841 caataaaccg ggcgacctca gccatccctt cctgattttc cgctttccag cgttcggcac
      901 gcagacgacg ggcttcattc tgcatggttg tgcttaccag accggagata ttgacatcat
      961 atatgccttg agcaactgat agctgtcgct gtcaactgtc actgtaatac gctgcttcat
     1021 agcatacctc tttttgacat acttcgggta tacatatcag tatatattct tataccgcaa
     1081 aaatcagcgc gcaaatacgc atactgttat ctggctttta gtaagccgga tccacgcgtt
     1141 tacgccccgc cctgccactc atcgcagtac tgttgtaatt cattaagcat tctgccgaca
     1201 tggaagccat cacaaacggc atgatgaacc tgaatcgcca gcggcatcag caccttgtcg
     1261 ccttgcgtat aatatttgcc catggtgaaa acgggggcga agaagttgtc catattggcc
     1321 acgtttaaat caaaactggt gaaactcacc cagggattgg ctgagacgaa aaacatattc
     1381 tcaataaacc ctttagggaa ataggccagg ttttcaccgt aacacgccac atcttgcgaa
     1441 tatatgtgta gaaactgccg gaaatcgtcg tggtattcac tccagagcga tgaaaacgtt
     1501 tcagtttgct catggaaaac ggtgtaacaa gggtgaacac tatcccatat caccagctca
     1561 ccgtctttca ttgccatacg gaattccgga tgagcattca tcaggcgggc aagaatgtga
     1621 ataaaggccg gataaaactt gtgcttattt ttctttacgg tctttaaaaa ggccgtaata
     1681 tccagctgaa cggtctggtt ataggtacat tgagcaactg actgaaatgc ctcaaaatgt
     1741 tctttacgat gccattggga tatatcaacg gtggtatatc cagtgatttt tttctccatt
     1801 ttagcttcct tagctcctga aaatctcgac ggatcctaac tcaaaatcca cacattatac
     1861 gagccggaag cataaagtgt aaagcctggg gtgcctaatg cggccgccat agtgactgga
     1921 tatgttgtgt tttacagtat tatgtagtct gttttttatg caaaatctaa tttaatatat
     1981 tgatatttat atcattttac gtttctcgtt cagctttttt gtacaaactt gtgatatcga
     2041 attcgctagc tacctttctc ttcttttttg gatctacctt tctcttcttt tttggggatc
     2101 cactagttca gctcttctgc ataataacta gaacaaaaac tcatctcaga agaggatctg
     2161 aatagcgccg tcgaccatca tcatcatcat cattgagttt aaacggtctc cagcttggct
     2221 gttttggcgg atgagagaag attttcagcc tgatacagat taaatcagaa cgcagaagcg
     2281 gtctgataaa acagaatttg cctggcggca gtagcgcggt ggtcccacct gaccccatgc
     2341 cgaactcaga agtgaaacgc cgtagcgccg atggtagtgt ggggtctccc catgcgagag
     2401 tagggaactg ccaggcatca aataaaacga aaggctcagt cgaaagactg ggcctttcgt
     2461 tttatctgtt gtttgtcggt gaacgctctc ctgagtagga caaatccgcc gggagcggat
     2521 ttgaacgttg cgaagcaacg gcccggaggg tggcgggcag gacgcccgcc ataaactgcc
     2581 aggcatcaaa ttaagcagaa ggccatcctg acggatggcc tttttgcgtt tctacaaact
     2641 cttttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     2701 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc
     2761 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     2821 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct
     2881 caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac
     2941 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtgtt gacgccgggc aagagcaact
     3001 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa
     3061 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga
     3121 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt
     3181 tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga
     3241 agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg
     3301 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat
     3361 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat
     3421 tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc
     3481 agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga
     3541 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc
     3601 agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     3661 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     3721 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     3781 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     3841 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     3901 accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     3961 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     4021 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     4081 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     4141 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     4201 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     4261 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     4321 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     4381 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc
     4441 tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac
     4501 cgagcgcagc gagtcagtga gcgaggaagc ggaagtgccc ctgatgcggt attttctcct
     4561 tacgcatctg tgcggtattt cacaccgcat atggtgcact ctcagtacaa tctgctctga
     4621 tgccgcatag ttaagccagt atacactccg ctatcgctac gtgactgggt catggctgcg
     4681 ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc
     4741 gcttacagac aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca
     4801 tcaccgaaac gcgcgaggca gcagatcaat tcgcgcgcga aggcgaagcg gcatgcataa
     4861 tgtgcctgtc aaatggacga agcagggatt ctgcaaaccc tatgctactc cgtcaagccg
     4921 tcaattgtct gattcgttac caattatgac aacttgacgg ctacatcatt cactttttct
     4981 tcacaaccgg cacggaactc gctcgggctg gccccggtgc attttttaaa tacccgcgag
     5041 aaatagagtt gatcgtcaaa accaacattg cgaccgacgg tggcgatagg catccgggtg
     5101 gtgctcaaaa gcagcttcgc ctggctgata cgttggtcct cgcgccagct taagacgcta
     5161 atccctaact gctggcggaa aagatgtgac agacgcgacg gcgacaagca aacatgctgt
     5221 gcgacgctgg cgatatcaaa attgctgtct gccaggtgat cgctgatgta ctgacaagcc
     5281 tcgcgtaccc gattatccat cggtggatgg agcgactcgt taatcgcttc catgcgccgc
     5341 agtaacaatt gctcaagcag atttatcgcc agcagctccg aatagcgccc ttccccttgc
     5401 ccggcgttaa tgatttgccc aaacaggtcg ctgaaatgcg gctggtgcgc ttcatccggg
     5461 cgaaagaacc ccgtattggc aaatattgac ggccagttaa gccattcatg ccagtaggcg
     5521 cgcggacgaa agtaaaccca ctggtgatac cattcgcgag cctccggatg acgaccgtag
     5581 tgatgaatct ctcctggcgg gaacagcaaa atatcacccg gtcggcaaac aaattctcgt
     5641 ccctgatttt tcaccacccc ctgaccgcga atggtgagat tgagaatata acctttcatt
     5701 cccagcggtc ggtcgataaa aaaatcgaga taaccgttgg cctcaatcgg cgttaaaccc
     5761 gccaccagat gggcattaaa cgagtatccc ggcagcaggg gatcattttg cgcttcagcc
     5821 atacttttca tactcccgcc attcagag
//
