LOCUS       sequence-175706-        5211 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  bacterial co-expression of folA (DHFR) and thyA(TS) with a barcode 
            within the noncoding region between the genes .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5211)
  AUTHORS   Schober AF, Mathis AD, Ingle C, Park JO, Chen L, Rabinowitz JD, 
            Junier I, Rivoire O, Reynolds KA
  TITLE     A Two-Enzyme Adaptive Unit within Bacterial Folate Metabolism.
  JOURNAL   Cell Rep. 2019 Jun 11;27(11):3359-3370.e7. doi: 
            10.1016/j.celrep.2019.05.030.
  PUBMED    31189117
REFERENCE   2  (bases 1 to 5211)
  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: 
            "10.1016/j.celrep.2019.05"; journalName: "Cell Rep"; date: 
            "2019-06-11- 11"; volume: "27"; issue: "11"; pages: "3359-3370.e7"
COMMENT     Sequence Label: pACYCDuet_G121VDHFR_R166QTS
FEATURES             Location/Qualifiers
     source          1..5211
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             12..56
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(94..112)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      108..155
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(375..1034)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     949..968
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(1035..1137)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(1529..1546)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1663..2207)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(1743..1762)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2432..3514)
                     /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     3476..3495
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3515..3592)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        3638..3656
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3657..3681
                     /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)."
     CDS             3737..3754
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             3767..4246
                     /codon_start=1
                     /gene="folA"
                     /product="E. coli dihydrofolate reductase"
                     /label=ecDHFR
                     /translation="MISLIAALAVDRVIGMENAMPWNLPADLAWFKRNTLNKPVIMGRH
                     TWESIGRPLPGRKNIILSSQPGTDDRVTWVKSVDEAIAACGDVPEIMVIGGGRVYEQFL
                     PKAQKLYLTHIDAEVEVDTHFPDYEPDDWESVFSEFHDADAQNSHSYCFEILERR"
     promoter        4319..4337
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    4338..4362
                     /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)."
ORIGIN
        1 tcgagtctgg taaagaaacc gctgctgcga aatttgaacg ccagcacatg gactcgtcta
       61 ctagcgcagc ttaattaacc taggctgctg ccaccgctga gcaataacta gcataacccc
      121 ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa acctcaggca tttgagaagc
      181 acacggtcac actgcttccg gtagtcaata aaccggtaaa ccagcaatag acataagcgg
      241 ctatttaacg accctgccct gaaccgacga ccgggtcgaa tttgctttcg aatttctgcc
      301 attcatccgc ttattatcac ttattcaggc gtagcaccag gcgtttaagg gcaccaataa
      361 ctgccttaaa aaaattacgc cccgccctgc cactcatcgc agtactgttg taattcatta
      421 agcattctgc cgacatggaa gccatcacag acggcatgat gaacctgaat cgccagcggc
      481 atcagcacct tgtcgccttg cgtataatat ttgcccatag tgaaaacggg ggcgaagaag
      541 ttgtccatat tggccacgtt taaatcaaaa ctggtgaaac tcacccaggg attggctgag
      601 acgaaaaaca tattctcaat aaacccttta gggaaatagg ccaggttttc accgtaacac
      661 gccacatctt gcgaatatat gtgtagaaac tgccggaaat cgtcgtggta ttcactccag
      721 agcgatgaaa acgtttcagt ttgctcatgg aaaacggtgt aacaagggtg aacactatcc
      781 catatcacca gctcaccgtc tttcattgcc atacggaact ccggatgagc attcatcagg
      841 cgggcaagaa tgtgaataaa ggccggataa aacttgtgct tatttttctt tacggtcttt
      901 aaaaaggccg taatatccag ctgaacggtc tggttatagg tacattgagc aactgactga
      961 aatgcctcaa aatgttcttt acgatgccat tgggatatat caacggtggt atatccagtg
     1021 atttttttct ccattttagc ttccttagct cctgaaaatc tcgataactc aaaaaatacg
     1081 cccggtagtg atcttatttc attatggtga aagttggaac ctcttacgtg ccgatcaacg
     1141 tctcattttc gccaaaagtt ggcccagggc ttcccggtat caacagggac accaggattt
     1201 atttattctg cgaagtgatc ttccgtcaca ggtatttatt cggcgcaaag tgcgtcgggt
     1261 gatgctgcca acttactgat ttagtgtatg atggtgtttt tgaggtgctc cagtggcttc
     1321 tgtttctatc agctgtccct cctgttcagc tactgacggg gtggtgcgta acggcaaaag
     1381 caccgccgga catcagcgct agcggagtgt atactggctt actatgttgg cactgatgag
     1441 ggtgtcagtg aagtgcttca tgtggcagga gaaaaaaggc tgcaccggtg cgtcagcaga
     1501 atatgtgata caggatatat tccgcttcct cgctcactga ctcgctacgc tcggtcgttc
     1561 gactgcggcg agcggaaatg gcttacgaac ggggcggaga tttcctggaa gatgccagga
     1621 agatacttaa cagggaagtg agagggccgc ggcaaagccg tttttccata ggctccgccc
     1681 ccctgacaag catcacgaaa tctgacgctc aaatcagtgg tggcgaaacc cgacaggact
     1741 ataaagatac caggcgtttc ccctggcggc tccctcgtgc gctctcctgt tcctgccttt
     1801 cggtttaccg gtgtcattcc gctgttatgg ccgcgtttgt ctcattccac gcctgacact
     1861 cagttccggg taggcagttc gctccaagct ggactgtatg cacgaacccc ccgttcagtc
     1921 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggaaa gacatgcaaa
     1981 agcaccactg gcagcagcca ctggtaattg atttagagga gttagtcttg aagtcatgcg
     2041 ccggttaagg ctaaactgaa aggacaagtt ttggtgactg cgctcctcca agccagttac
     2101 ctcggttcaa agagttggta gctcagagaa ccttcgaaaa accgccctgc aaggcggttt
     2161 tttcgttttc agagcaagag attacgcgca gaccaaaacg atctcaagaa gatcatctta
     2221 ttaatcagat aaaatatttc tagatttcag tgcaatttat ctcttcaaat gtagcacctg
     2281 aagtcagccc catacgatat aagttgtaat tctcatgtta gtcatgcccc gcgcccaccg
     2341 gaaggagctg actgggttga aggctctcaa gggcatcggt cgagatcccg gtgcctaatg
     2401 agtgagctaa cttacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     2461 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     2521 gcgccagggt ggtttttctt ttcaccagtg agacgggcaa cagctgattg cccttcaccg
     2581 cctggccctg agagagttgc agcaagcggt ccacgctggt ttgccccagc aggcgaaaat
     2641 cctgtttgat ggtggttaac ggcgggatat aacatgagct gtcttcggta tcgtcgtatc
     2701 ccactaccga gatgtccgca ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc
     2761 ccagcgccat ctgatcgttg gcaaccagca tcgcagtggg aacgatgccc tcattcagca
     2821 tttgcatggt ttgttgaaaa ccggacatgg cactccagtc gccttcccgt tccgctatcg
     2881 gctgaatttg attgcgagtg agatatttat gccagccagc cagacgcaga cgcgccgaga
     2941 cagaacttaa tgggcccgct aacagcgcga tttgctggtg acccaatgcg accagatgct
     3001 ccacgcccag tcgcgtaccg tcttcatggg agaaaataat actgttgatg ggtgtctggt
     3061 cagagacatc aagaaataac gccggaacat tagtgcaggc agcttccaca gcaatggcat
     3121 cctggtcatc cagcggatag ttaatgatca gcccactgac gcgttgcgcg agaagattgt
     3181 gcaccgccgc tttacaggct tcgacgccgc ttcgttctac catcgacacc accacgctgg
     3241 cacccagttg atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg
     3301 ccagactgga ggtggcaacg ccaatcagca acgactgttt gcccgccagt tgttgtgcca
     3361 cgcggttggg aatgtaattc agctccgcca tcgccgcttc cactttttcc cgcgttttcg
     3421 cagaaacgtg gctggcctgg ttcaccacgc gggaaacggt ctgataagag acaccggcat
     3481 actctgcgac atcgtataac gttactggtt tcacattcac caccctgaat tgactctctt
     3541 ccgggcgcta tcatgccata ccgcgaaagg ttttgcgcca ttcgatggtg tccgggatct
     3601 cgacgctctc ccttatgcga ctcctgcatt aggaaattaa tacgactcac tataggggaa
     3661 ttgtgagcgg ataacaattc ccctgtagaa ataattttgt ttaactttaa taaggagata
     3721 taccatgggc agcagccatc accatcatca ccacagccag gatccgatga tcagtctgat
     3781 tgcggcgtta gcggtagatc gcgttatcgg catggaaaac gccatgccgt ggaacctgcc
     3841 tgccgatctc gcctggttta aacgcaacac cttaaataaa cccgtgatta tgggccgcca
     3901 tacctgggaa tcaatcggtc gtccgttgcc aggacgcaaa aatattatcc tcagcagtca
     3961 accgggtacg gacgatcgcg taacgtgggt gaagtcggtg gatgaagcca tcgcggcgtg
     4021 tggtgacgta ccagaaatca tggtgattgg cggcggtcgc gtttatgaac agttcttgcc
     4081 aaaagcgcaa aaactgtatc tgacgcatat cgacgcagaa gtggaagtcg acacccattt
     4141 cccggattac gagccggatg actgggaatc ggtattcagc gaattccacg atgctgatgc
     4201 gcagaactct cacagctatt gctttgagat tctggagcgg cggtaacata agcggccgcg
     4261 acagatgctt aagtcgaaca gaaagtaatc gtattgtaca cggccgcata atcgaaatta
     4321 atacgactca ctatagggga attgtgagcg gataacaatt ccccatctta gtatattagt
     4381 taagtataag aaggagatat acatatggca gatctatgaa acagtacctg gagctgatgc
     4441 aaaaagttct ggatgagggt acccagaaaa acgaccgcac gggtaccgga acgctgagca
     4501 tttttggcca tcagatgcgt tttaacctgc aggacggatt cccgctggtt acgaccaaac
     4561 gctgccacct gcgtagcatt attcatgagc tgctgtggtt tctgcaaggt gacactaaca
     4621 tcgcgtatct gcacgaaaac aatgtgacga tctgggatga atgggccgat gaaaacggcg
     4681 atctgggccc agtgtatggt aaacagtggc gcgcctggcc aacgccagat ggccgccaca
     4741 tcgatcagat cacgaccgtg ctgaatcaac tgaaaaacga cccggacagc cgccgcatta
     4801 ttgtttccgc gtggaatgtg ggtgaactgg ataaaatggc gctggcgccg tgccatgcat
     4861 tcttccagtt ttacgttgcg gacggtaagc tgagctgtca actttatcag cagagctgcg
     4921 atgttttcct cggcctgccg ttcaatatcg ccagctacgc gttactggtg catatgatgg
     4981 cgcagcagtg cgacctggag gtgggtgatt ttgtctggac cggcggtgat acccacctgt
     5041 acagcaacca tatggaccaa acgcatctgc aattgagccg tgagccgcgt ccgctgccga
     5101 aactgattat taagcgcaaa cccgagagca tctttgacta tcgtttcgaa gacttcgaga
     5161 ttgaaggcta cgatccgcac ccgggtatca aagcgccggt ggctatctaa c
//
