LOCUS       sequence-338830-        7004 bp    DNA     circular SYN 15-APR-2025
DEFINITION  pylBCD variant in pBAD backbone.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7004)
  AUTHORS   Ho JML, Miller CA, Smith KA, Mattia JR, Bennett MR
  TITLE     Improved pyrrolysine biosynthesis through phage assisted 
            non-continuous directed evolution of the complete pathway.
  JOURNAL   Nat Commun. 2021 Jun 24;12(1):3914. doi: 10.1038/s41467-021-24183-9.
  PUBMED    34168131
REFERENCE   2  (bases 1 to 7004)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2021-06-24"; volume: "12(1):3914. doi"; pages: " 
            10.1038/s41467-021-24183-9"
COMMENT     Sequence Label: pJH573
FEATURES             Location/Qualifiers
     source          1..7004
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      28..55
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        74..165
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(166..234,235..1026)
                     /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(384..403)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1197..1785
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1686..1705
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1950..1967
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    1982..2122
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2048..2067)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2208..2230
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2386..3264)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        3291..3575
                     /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     3498..3517
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     primer_bind     complement(6684..6701)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      6854..6940
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 cataaactgc caggcatcaa attaagcaga aggccatcct gacggatggc ctttttgcgt
       61 ttctacaaac tcttttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
      121 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
      181 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
      241 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
      301 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
      361 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtgt tgacgccggg
      421 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
      481 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
      541 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
      601 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
      661 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
      721 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
      781 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
      841 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
      901 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
      961 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     1021 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     1081 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     1141 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     1201 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     1261 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     1321 agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag
     1381 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     1441 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     1501 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     1561 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     1621 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     1681 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     1741 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     1801 gcctttttac ggttcctggc cttttgctgg ccttttgctg gccttttgct cacatgttct
     1861 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     1921 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     1981 gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atatggtgca
     2041 ctctcagtac aatctgctct gatgccgcat agttaagcca gtatacactc cgctatcgct
     2101 acgtgactgg gtcatggctg cgccccgaca cccgccaaca cccgctgacg cgccctgacg
     2161 ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat
     2221 gtgtcagagg ttttcaccgt catcaccgaa acgcgcgagg cagcagatca attcgcgcgc
     2281 gaaggcgaag cggcatgcat aatgtgcctg tcaaatggac gaagcaggga ttctgcaaac
     2341 cctatgctac tccgtcaagc cgtcaattgt ctgattcgtt accaattatg acaacttgac
     2401 ggctacatca ttcacttttt cttcacaacc ggcacggaac tcgctcgggc tggccccggt
     2461 gcatttttta aatacccgcg agaaatagag ttgatcgtca aaaccaacat tgcgaccgac
     2521 ggtggcgata ggcatccggg tggtgctcaa aagcagcttc gcctggctga tacgttggtc
     2581 ctcgcgccag cttaagacgc taatccctaa ctgctggcgg aaaagatgtg acagacgcga
     2641 cggcgacaag caaacatgct gtgcgacgct ggcgatatca aaattgctgt ctgccaggtg
     2701 atcgctgatg tactgacaag cctcgcgtac ccgattatcc atcggtggat ggagcgactc
     2761 gttaatcgct tccatgcgcc gcagtaacaa ttgctcaagc agatttatcg ccagcagctc
     2821 cgaatagcgc ccttcccctt gcccggcgtt aatgatttgc ccaaacaggt cgctgaaatg
     2881 cggctggtgc gcttcatccg ggcgaaagaa ccccgtattg gcaaatattg acggccagtt
     2941 aagccattca tgccagtagg cgcgcggacg aaagtaaacc cactggtgat accattcgcg
     3001 agcctccgga tgacgaccgt agtgatgaat ctctcctggc gggaacagca aaatatcacc
     3061 cggtcggcaa acaaattctc gtccctgatt tttcaccacc ccctgaccgc gaatggtgag
     3121 attgagaata taacctttca ttcccagcgg tcggtcgata aaaaaatcga gataaccgtt
     3181 ggcctcaatc ggcgttaaac ccgccaccag atgggcatta aacgagtatc ccggcagcag
     3241 gggatcattt tgcgcttcag ccatactttt catactcccg ccattcagag aagaaaccaa
     3301 ttgtccatat tgcatcagac attgccgtca ctgcgtcttt tactggctct tctcgctaac
     3361 caaaccggta accccgctta ttaaaagcat tctgtaacaa agcgggacca aagccatgac
     3421 aaaaacgcgt aacaaaagtg tctataatca cggcagaaaa gtccacattg attatttgca
     3481 cggcgtcaca ctttgctatg ccatagcatt tttatccata agattagcgg atcctacctg
     3541 acgcttttta tcgcaactct ctactgtttc tccatacccg ttttttgggc taacaggagg
     3601 aattaaccat gattaagaag atggcgactg aagatctgga tcgttttggc gaaaaaatta
     3661 ttgaaggatt tcagttatct gacgacgatt tacgcgcttt gcttagcctg gaatcggaag
     3721 aagaattgga aaagctgtat tacgtcgcac gtaaagtacg caaccattac tttggaaatc
     3781 gcgtattcct gaactgtttt atctacttca gcacatactg caaaaaccaa tgtgcgtttt
     3841 gctattataa ctgcaaaaac gagattaacc gctatcgttt gtcccgcgaa gaaatcaaag
     3901 agacttgcaa ggcccttaaa ggtgcgggtt tccatatgat tgatctgact atgggggagg
     3961 atccctatta ttacgaggaa cctgatcgct tcgtggagct ggtccaaatg gttaagaatg
     4021 agcttggact tccaatcatg atttcgccag gagtaatgga taatgctacc cttttaaaag
     4081 cacgcgaaaa gggagctaat ttcttcgcct tatatcaaga aacctatgat caagagttgt
     4141 acgagaagtt gcgcgtaggt cagtcgttca aggggcgcta caatactcgt cgttttgcta
     4201 aagaacaagg gtattgcatt gaagacggta tcttaacggg agtcgggaac gatatcgagt
     4261 caacgatcaa atctttacgc ggcatgaaat caaatgatcc cgacatggtt cgcgtgatga
     4321 ctttcttacc ccaggaagga actccccttg agcgttttaa ggaaaatagt aacctgagcg
     4381 aactgaaaat cattgcgatc ttgcgtctga tctttccgaa gtgcttaatc cccgcgtcgt
     4441 tggacctgga agggatcgac gggatggtac accgtcttaa cgcaggtgct aatatcgtga
     4501 ctagcatcct tcctagtgat tcaaaacttg agggagttgc taactacgat cgtgacctgg
     4561 aagagcgcga ccgcgacatt aaaagcgtca tcaagcgcct tgagggaatg ggcatggaac
     4621 cagcgccaca ggcggaattc gagactgtac tgcgctgcta acatgaaaac catctgcctg
     4681 attggtggta aattgcaggg ttttgaggcc gcgtatctgt ctaaaaaagc cgatatgaaa
     4741 gtagtggtta tcgacaagaa cccccaagct cttatccgca attacgcaga cgagtttcat
     4801 tgttttgatg ttatcaagga accagagaaa cttcttgaaa tcagcaaaaa tgtagacgct
     4861 atcttgcccg tcaacgaaaa cttagaatgt attaaattct taagtagtgt aaaagaggaa
     4921 ttttcttgcc cagtactgtt cgattttgag gcttaccgca tctcgcgcga taagaagaaa
     4981 tcgaaagagt atttccgttc catcggcatt ccgacaccgc aagacaaacc tggtcgtcct
     5041 ccatacttcg ttaagccacc gtgcgaaagc tcgtcggttg gtgctcgcat tatctacgat
     5101 gaagaagaac tgggagaatt ggagcccggc atgctggtag aagagtatgt ggaaggtgag
     5161 gttatttcac ttgaagtaat cggggacggt acccactttg cagttgttaa agagacactt
     5221 attcacatcg accgcaccta tgactgccat atggtaactc ccctgccatc tgatttgtct
     5281 ttccgtgaga tcagctattc tttagcggcc aacctgcctt tgaaggggat catggacgtt
     5341 gaagcaattt cgggcccgca aggtttgaag gtcattgaaa ttgattcacg tttcccctcc
     5401 cagaccccca cagccgtgta ttattcgtcg ggagtgaatt taattgagct gttgtttcgt
     5461 gcctttggag aaggggtgga ggaagttaaa acccttccag aggaccgcta ctgtatctac
     5521 gagcacctga tgcttgcaga aaatggggcc ctgatcccag taggtgagca ggttctgtcg
     5581 atggggaatg actatgggaa atactatgaa gaaccaggaa ttgaaatctt tttatgtcgc
     5641 ggagaggacc ccgtgtttac gcttgtcttt tggggaaagg accgtgagga agccgaaaac
     5701 aaaaagcgca ctggtttgct gattttgaag gaccgcttcg gggcagccgt ttaaaatggc
     5761 aaaaatcgga taagaataaa aatttgccca aagtaaaatt tgtttaaaac agaaaaagga
     5821 gataaaaaat ggctttgttg actcccgagg acctggagaa cattaaccgt caacttcacg
     5881 aagccgattc tacggtacgt cgtgtgactg ggttagatat caagggagtt tgtaaagatt
     5941 tctatgggac tactccatgc tgtgagaagg tcggtatcgt cccggtaact agtggcaacg
     6001 gcatcattgg caacttttct gaaagtcttc gtgcgatcac tgagtacttt ggatttgact
     6061 cattcgtgac gaccatgcca gacgttagtg gctactatga ggcggtccgt aatggagcac
     6121 aaattattct tatggcagat gataatacgt ttcttgcaca caatttaaag aacggcaaga
     6181 tcgctaataa tcaaccgtgt accggaatta tctacgcgga gatcgcctcg cgctatatga
     6241 aagccgattc caaagatgtg cttgccgtcg ggcttggaaa agtggggttt ccaggcgctg
     6301 cacatttggt taataagggg tttcgtgtgt atgggtacga cgcggataag gcgttgttag
     6361 aaaagactgt atcagacctt gggatcacac ctttcgagcc tgagaatccc cgtcgcttct
     6421 caatcatttt tgaggcgacc ccgtgtgctg atacagtacc agagtctgtc atcagcgaca
     6481 attgtgtcat ctctacgccc ggtatccctt gcgcgatcag ccgtgaactt caagaaaaat
     6541 acggggtgga gttgattatg gaacctctgg gtattggaac tgcttctatg ctgtatagca
     6601 ttttgtaaag ctcgagatct gcagctggta ccatatggga attcgaagct tggctgtttt
     6661 ggcggatgag agaagatttt cagcctgata cagattaaat cagaacgcag aagcggtctg
     6721 ataaaacaga atttgcctgg cggcagtagc gcggtggtcc cacctgaccc catgccgaac
     6781 tcagaagtga aacgccgtag cgccgatggt agtgtggggt ctccccatgc gagagtaggg
     6841 aactgccagg catcaaataa aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat
     6901 ctgttgtttg tcggtgaacg ctctcctgag taggacaaat ccgccgggag cggatttgaa
     6961 cgttgcgaag caacggcccg gagggtggcg ggcaggacgc ccgc
//
