LOCUS       sequence-213174-        6665 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Phe-tRNA synthetase A.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6665)
  AUTHORS   Villarreal F, Contreras-Llano LE, Chavez M, Ding Y, Fan J, Pan T, 
            Tan C
  TITLE     Synthetic microbial consortia enable rapid assembly of pure 
            translation machinery.
  JOURNAL   Nat Chem Biol. 2018 Jan;14(1):29-35. doi: 10.1038/nchembio.2514. 
            Epub 2017 Nov 13.
  PUBMED    29131146
REFERENCE   2  (bases 1 to 6665)
  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"; doi: 
            "10.1038/nchembio.2514"; journalName: "Nat Chem Biol"; date: 
            "2018-01"; volume: "14"; issue: "1"; pages: "29-35"
COMMENT     Sequence Label: B6-4
FEATURES             Location/Qualifiers
     source          1..6665
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     106..125
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     209..228
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        209..227
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    228..252
                     /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)."
     RBS             267..289
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             299..316
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1374..1392)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1388..1435
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(1610..1615,1616..1632,1633..1638))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(1665..1683)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1751..1855
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1856..1924,1925..2716)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2074..2093)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2887..3475
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3376..3395
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3629..3646
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3661..3800
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3726..3745)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3886..3908
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3902..4093)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5405..6487)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     6449..6468
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6488..6565)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 caaggagatg gcgcccaaca gtcccccggc cacggggcct gccaccatac ccacgccgaa
       61 acaagcgctc atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat
      121 ataggcgcca gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta
      181 gaggatcgag atctcgatcc cgcgaaatta atacgactca ctatagggga attgtgagcg
      241 gataacaatt cccctctaga aataattttg tttaacttta agaaggagat ataccatgca
      301 tcatcatcac catcatttat cacatctcgc agaactggtt gccagtgcga aggcggccat
      361 tagccaggcg tcagatgttg ccgcgttaga taatgtgcgc gtcgaatatt tgggtaaaaa
      421 agggcactta acccttcaga tgacgaccct gcgtgagctg ccgccagaag agcgtccggc
      481 agctggtgcg gttatcaacg aagcgaaaga gcaggttcag caggcgctga atgcgcgtaa
      541 agcggaactg gaaagctctg cactgaatgc gcgtctggcg gcggaaacga ttgatgtctc
      601 tctgccaggt cgtcgcattg aaaacggcgg tctgcatccg gttacccgta ccatcgaccg
      661 tatcgaaagt ttcttcggtg agcttggctt taccgtggca accgggccgg aaatcgaaga
      721 cgattatcat aacttcgatg ctctgaacat tcctggtcac cacccggcgc gcgctgacca
      781 cgacactttc tggtttgaca ctacccgcct gctgcgtacc cagacctctg gcgtacagat
      841 ccgcaccatg aaagcccagc agccaccgat tcgtatcatc gcgcctggcc gtgtttatcg
      901 taacgactac gaccagactc acacgccgat gttccatcag atggaaggtc tgattgttga
      961 taccaacatc agctttacca acctgaaagg cacgctgcac gacttcctgc gtaacttctt
     1021 tgaggaagat ttgcagattc gcttccgtcc ttcctacttc ccgtttaccg aaccttctgc
     1081 agaagtggac gtcatgggta aaaacggtaa atggctggaa gtgctgggct gcgggatggt
     1141 gcatccgaac gtgttgcgta acgttggcat cgacccggaa gtttactctg gtttcgcctt
     1201 cgggatgggg atggagcgtc tgactatgtt gcgttacggc gtcaccgacc tgcgttcatt
     1261 cttcgaaaac gatctgcgtt tcctcaaaca gtttaaatat taaagcagag gccatatgct
     1321 cgaggatccg gctgctaaca aagcccgaaa ggaagctgag ttggctgctg ccaccgctga
     1381 gcaataacta gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa
     1441 aggaggaact atatccggat atcccgcaag aggcccggca gtaccggcat aaccaagcct
     1501 atgcctacag catccagggt gacggtgccg aggatgacga tgagcgcatt gttagatttc
     1561 atacacggtg cctgactgcg ttagcaattt aactgtgata aactaccgca ttaaagctta
     1621 tcgatgataa gctgtcaaac atgagaattc ttgaagacga aagggcctcg tgatacgcct
     1681 atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg
     1741 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     1801 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
     1861 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     1921 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     1981 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     2041 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtgt
     2101 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     2161 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     2221 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     2281 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     2341 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgc
     2401 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     2461 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     2521 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     2581 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     2641 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     2701 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     2761 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     2821 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     2881 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     2941 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     3001 tggcttcagc agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca
     3061 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     3121 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     3181 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     3241 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     3301 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     3361 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     3421 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     3481 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     3541 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac
     3601 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg
     3661 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca atggtgcact
     3721 ctcagtacaa tctgctctga tgccgcatag ttaagccagt atacactccg ctatcgctac
     3781 gtgactgggt catggctgcg ccccgacacc cgccaacacc cgctgacgcg ccctgacggg
     3841 cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt
     3901 gtcagaggtt ttcaccgtca tcaccgaaac gcgcgaggca gctgcggtaa agctcatcag
     3961 cgtggtcgtg aagcgattca cagatgtctg cctgttcatc cgcgtccagc tcgttgagtt
     4021 tctccagaag cgttaatgtc tggcttctga taaagcgggc catgttaagg gcggtttttt
     4081 cctgtttggt cactgatgcc tccgtgtaag ggggatttct gttcatgggg gtaatgatac
     4141 cgatgaaacg agagaggatg ctcacgatac gggttactga tgatgaacat gcccggttac
     4201 tggaacgttg tgagggtaaa caactggcgg tatggatgcg gcgggaccag agaaaaatca
     4261 ctcagggtca atgccagcgc ttcgttaata cagatgtagg tgttccacag ggtagccagc
     4321 agcatcctgc gatgcagatc cggaacataa tggtgcaggg cgctgacttc cgcgtttcca
     4381 gactttacga aacacggaaa ccgaagacca ttcatgttgt tgctcaggtc gcagacgttt
     4441 tgcagcagca gtcgcttcac gttcgctcgc gtatcggtga ttcattctgc taaccagtaa
     4501 ggcaaccccg ccagcctagc cgggtcctca acgacaggag cacgatcatg cgcacccgtg
     4561 gccaggaccc aacgctgccc gagatgcgcc gcgtgcggct gctggagatg gcggacgcga
     4621 tggatatgtt ctgccaaggg ttggtttgcg cattcacagt tctccgcaag aattgattgg
     4681 ctccaattct tggagtggtg aatccgttag cgaggtgccg ccggcttcca ttcaggtcga
     4741 ggtggcccgg ctccatgcac cgcgacgcaa cgcggggagg cagacaaggt atagggcggc
     4801 gcctacaatc catgccaacc cgttccatgt gctcgccgag gcggcataaa tcgccgtgac
     4861 gatcagcggt ccaatgatcg aagttaggct ggtaagagcc gcgagcgatc cttgaagctg
     4921 tccctgatgg tcgtcatcta cctgcctgga cagcatggcc tgcaacgcgg gcatcccgat
     4981 gccgccggaa gcgagaagaa tcataatggg gaaggccatc cagcctcgcg tcgcgaacgc
     5041 cagcaagacg tagcccagcg cgtcggccgc catgccggcg ataatggcct gcttctcgcc
     5101 gaaacgtttg gtggcgggac cagtgacgaa ggcttgagcg agggcgtgca agattccgaa
     5161 taccgcaagc gacaggccga tcatcgtcgc gctccagcga aagcggtcct cgccgaaaat
     5221 gacccagagc gctgccggca cctgtcctac gagttgcatg ataaagaaga cagtcataag
     5281 tgcggcgacg atagtcatgc cccgcgccca ccggaaggag ctgactgggt tgaaggctct
     5341 caagggcatc ggtcgagatc ccggtgccta atgagtgagc taacttacat taattgcgtt
     5401 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
     5461 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgccag ggtggttttt cttttcacca
     5521 gtgagacggg caacagctga ttgcccttca ccgcctggcc ctgagagagt tgcagcaagc
     5581 ggtccacgct ggtttgcccc agcaggcgaa aatcctgttt gatggtggtt aacggcggga
     5641 tataacatga gctgtcttcg gtatcgtcgt atcccactac cgagatatcc gcaccaacgc
     5701 gcagcccgga ctcggtaatg gcgcgcattg cgcccagcgc catctgatcg ttggcaacca
     5761 gcatcgcagt gggaacgatg ccctcattca gcatttgcat ggtttgttga aaaccggaca
     5821 tggcactcca gtcgccttcc cgttccgcta tcggctgaat ttgattgcga gtgagatatt
     5881 tatgccagcc agccagacgc agacgcgccg agacagaact taatgggccc gctaacagcg
     5941 cgatttgctg gtgacccaat gcgaccagat gctccacgcc cagtcgcgta ccgtcttcat
     6001 gggagaaaat aatactgttg atgggtgtct ggtcagagac atcaagaaat aacgccggaa
     6061 cattagtgca ggcagcttcc acagcaatgg catcctggtc atccagcgga tagttaatga
     6121 tcagcccact gacgcgttgc gcgagaagat tgtgcaccgc cgctttacag gcttcgacgc
     6181 cgcttcgttc taccatcgac accaccacgc tggcacccag ttgatcggcg cgagatttaa
     6241 tcgccgcgac aatttgcgac ggcgcgtgca gggccagact ggaggtggca acgccaatca
     6301 gcaacgactg tttgcccgcc agttgttgtg ccacgcggtt gggaatgtaa ttcagctccg
     6361 ccatcgccgc ttccactttt tcccgcgttt tcgcagaaac gtggctggcc tggttcacca
     6421 cgcgggaaac ggtctgataa gagacaccgg catactctgc gacatcgtat aacgttactg
     6481 gtttcacatt caccaccctg aattgactct cttccgggcg ctatcatgcc ataccgcgaa
     6541 aggttttgcg ccattcgatg gtgtccggga tctcgacgct ctcccttatg cgactcctgc
     6601 attaggaagc agcccagtag taggttgagg ccgttgagca ccgccgccgc aaggaatggt
     6661 gcatg
//
