LOCUS       sequence-201203-        4328 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Formaldehyde biosensor plasmid with sGFP reporter and modified frm 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4328)
  AUTHORS   Woolston BM, Roth T, Kohale I, Liu D, Stephanopoulos G
  TITLE     Development of a Formaldehyde Biosensor with Application to 
            Synthetic Methylotrophy.
  JOURNAL   Biotechnol Bioeng. 2017 Sep 16. doi: 10.1002/bit.26455.
  PUBMED    28921510
REFERENCE   2  (bases 1 to 4328)
  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"; journalName: "Biotechnol 
            Bioeng. 2017 Sep 16. doi: 10.1002/bit.26455."
COMMENT     Sequence Label: pTR47m4-GFP
FEATURES             Location/Qualifiers
     source          1..4328
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(1..951)
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLRNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKHDLNGSFSWLTQKQRTTLENILAKYGRI"
     rep_origin      999..1221
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     CDS             join(1897..1899,1900..2613)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /note="mammalian codon-optimized"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     promoter        3058..3112
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
     protein_bind    3059..3077
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3089..3107
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O1 for the tetR and tetA genes"
     protein_bind    3113..3131
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             complement(join(3257..4048,4049..4117))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3880..3899
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4118..4222)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4290..4308
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcagatcctt ccgtatttag ccagtatgtt ctctagtgtg gttcgttgtt tttgcgtgag
       61 ccatgagaac gaaccattga gatcatgctt actttgcatg tcactcaaaa attttgcctc
      121 aaaactggtg agctgaattt ttgcagttaa agcatcgtgt agtgtttttc ttagtccgtt
      181 acgtaggtag gaatctgatg taatggttgt tggtattttg tcaccattca tttttatctg
      241 gttgttctca agttcggtta cgagatccat ttgtctatct agttcaactt ggaaaatcaa
      301 cgtatcagtc gggcggcctc gcttatcaac caccaatttc atattgctgt aagtgtttaa
      361 atctttactt attggtttca aaacccattg gttaagcctt ttaaactcat ggtagttatt
      421 ttcaagcatt aacatgaact taaattcatc aaggctaatc tctatatttg ccttgtgagt
      481 tttcttttgt gttagttctt ttaataacca ctcataaatc ctcatagagt atttgttttc
      541 aaaagactta acatgttcca gattatattt tatgaatttt tttaactgga aaagataagg
      601 caatatctct tcactaaaaa ctaattctaa tttttcgctt gagaacttgg catagtttgt
      661 ccactggaaa atctcaaagc ctttaaccaa aggattcctg atttccacag ttctcgtcat
      721 cagctctctg gttgctttag ctaatacacc ataagcattt tccctactga tgttcatcat
      781 ctgagcgtat tggttataag tgaacgatac cgtccgttct ttccttgtag ggttttcaat
      841 cgtggggttg agtagtgcca cacagcataa aattagcttg gtttcatgct ccgttaagtc
      901 atagcgacta atcgctagtt catttgcttt gaaaacaact aattcagaca tacatctcaa
      961 ttggtctagg tgattttaat cactatacca attgagatgg gctagtcaat gataattact
     1021 agtccttttc ctttgagttg tgggtatctg taaattctgc tagacctttg ctggaaaact
     1081 tgtaaattct gctagaccct ctgtaaattc cgctagacct ttgtgtgttt tttttgttta
     1141 tattcaagtg gttataattt atagaataaa gaaagaataa aaaaagataa aaagaataga
     1201 tcccagccct gtgtataact cactacttta gtcagttccg cagtattaca aaaggatgtc
     1261 gcaaacgctg tttgctcctc tacaaaacag accttaaaac cctaaaggct taagtagcac
     1321 cctcgcaagc tcgggcaaat cgctgaatat tccttttgtc tccgaccatc aggcacctga
     1381 gtcgctgtct ttttcgtgac attcagttcg ctgcgctcac ggctctggca gtgaatgggg
     1441 gtaaatggca ctacaggcgc cttttatgga ttcatgcaag gaaactaccc ataatacaag
     1501 aaaagcccgt cacgggcttc tcagggcgtt ttatggcggg tctgctatgt ggtgctatct
     1561 gactttttgc tgttcagcag ttcctgccct ctgattttcc agtctgacca cttcggatta
     1621 tcccgtgaca ggtcattcag actggctaat gcacccagta aggcagcggt atcatcaaca
     1681 ggcttacccg tcttactgtc aagaggacat ccggtcaaat aaaacgaaag gctcagtcga
     1741 aagactgggc ctttcgtttt gctgaggaga cttagggacc ctaaatttta ttctccagtg
     1801 ttatatacta taggggggta tgcattgaca tatagcatac actcctatag tatatgcgtg
     1861 cagataacct gcaggtgcag taaggaggaa aaaaaaatga gcaagggcga agagcttttt
     1921 accggtgttg tgccgatttt agtagaactg gacggagacg tgaacggtca taagttctct
     1981 gttcgtggcg aaggagaggg agatgccacc aatggtaagc tgaccctgaa gttcatctgt
     2041 accaccggta agctgcccgt gccttggccg acgctggtca caacgttgac gtatggcgtc
     2101 caatgctttt cacgctatcc agatcacatg aaacgccacg acttttttaa aagcgcaatg
     2161 cctgaaggtt atgtgcagga acggactatt agcttcaaag acgatgggac gtataagacc
     2221 cgcgcggaag tgaaatttga aggcgatacc ttagttaacc gcattgaatt aaaaggtatc
     2281 gatttcaaag aggatgggaa tatcctgggg cacaaattgg aatacaactt taattcgcac
     2341 aacgtataca ttacagcgga taaacagaaa aatggcatca aagccaactt taaaatccgt
     2401 cataacgtag aggacggttc cgtgcagctg gctgatcatt accagcagaa tactccgatt
     2461 ggcgatggcc ccgttctgct cccggataat cattacctgt ctacacaaag cgttcttagt
     2521 aaagacccaa acgagaagcg tgaccatatg gtcctgttgg aattcgtcac ggcagcgggg
     2581 attactcatg gcatggatga actctataag taaacggagc caatgtacgc aaaaaacccc
     2641 gcttcagcgg ggttttttcg ccaaaaaaaa ccccgccctg tcaggggcgg ggtttttttt
     2701 tagggcccta ctgactgtct atttaagata ggctcgaacc agttcaatag tgtcgtcaac
     2761 ggattggctg acttcgcggc tgtagcagtc atttcggtca aacgtttccc ggatatggct
     2821 ttcaagcact tctgccatca gcccattagc cgcgccccga acggcagcga tctgttggag
     2881 tatggcacgg cattcggcat caccctccag cgaccgttcc agagcatcaa tctgccctcg
     2941 aatacgacga actcgagtaa ggaccttttt cttctcttcc ggagtactgg gcattttttt
     3001 ttcctcctta taagggggta gggggtctag atttttgtcg aactattcat ttcacttttc
     3061 tctatcactg atagggagtg gtaaaataac tctatcaatg atagagtgtc aatctctatc
     3121 actgataggg atcatctggc cattcgatgg acttaattaa cggcactgcg aaaaaacccc
     3181 gccgaagcgg ggttttttgc gaaatataat gaccctcttg ataacccaag agggcatttt
     3241 ttaatgccca tggcgtttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     3301 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     3361 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     3421 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     3481 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     3541 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     3601 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     3661 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     3721 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     3781 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     3841 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     3901 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     3961 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     4021 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     4081 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     4141 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     4201 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa
     4261 ccattattat catgacatta acctataaaa ataggcgtat cacgaggccc ttaggccaaa
     4321 taggccgt
//
