LOCUS       sequence-335948-        6421 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes PmgtC-sfgfp reporter and TetR-controlled expression of S. 
            Typhimurium PhoP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6421)
  AUTHORS   Brink KR, Mu AM, Hoang KV, Groszman K, Gunn JS, Tabor JJ
  TITLE     High-throughput discovery of peptide activators of a bacterial 
            sensor kinase
  JOURNAL   bioRxiv
REFERENCE   2  (bases 1 to 6421)
  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: "bioRxiv"
COMMENT     Sequence Label: pAM003
FEATURES             Location/Qualifiers
     source          1..6421
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             445..456
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     primer_bind     complement(907..925)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        990..1094
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1095..1163,1164..1955)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFSVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLYRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1313..1332)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      1985..2079
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     rep_origin      2571..2793
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     CDS             2841..3791
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIMNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
     primer_bind     4053..4070
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(4154..4240)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(4246..4872)
                     /codon_start=1
                     /gene="tetR"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MMSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWH
                     VKNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGT
                     RPTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPT
                     TDSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     4777..4796
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        complement(4907..4941)
                     /label=BBa_J23104
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.72"
     promoter        complement(join(5803..5828,5829..5834,5835..5851,5852..5857,
                     5858..5876))
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    5833..5851
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5858..5876
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     terminator      complement(5953..5980)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      complement(5996..6067)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(join(6146..6421,1..438))
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
ORIGIN
        1 gttaaaattg tattccagct tatggcccag gatattgccg tcttctttaa agtcaatgcc
       61 tttcagctca atgcggttta ccagggtatc gccttcaaat ttcacttccg cacgcgtttt
      121 gtacgtgccg tcatccttaa aggaaatcgt gcgttcctgc acatagcctt ccggcatggc
      181 ggacttgaag aagtcatgct gcttcatatg gtccggataa cgagcaaagc actgaacacc
      241 ataagtcagc gtcgttacca gagtcggcca aggtaccggc agtttaccag tagtacagat
      301 gaacttcagc gtcagtttac cattagttgc gtcaccttca ccctcgccac gcacggaaaa
      361 cttatgaccg ttgacatcac catccagttc caccagaata gggacgacac cagtgaacag
      421 ctcttcgcct ttacgcatct agtatttctc ctcttttttt tctggaagaa taagtacgtg
      481 ctatatttag cacacgacaa atttctgccc tgaggcaatt agcgtcgtgt taacggcgga
      541 ggaaataaag cgtgtttaaa catgaacatg acgattacct cgcgacaatg cgcgtgttgt
      601 aacattattg ggatgacaat cgtcacaagg ggagattgct gcccaccggt tcggcaagca
      661 gcgcaggaat agcgtgcgtt aacttgccag atgatttacg taaaagtgac tgtccaacat
      721 attactcctg catgaatttg cgcttattat agtcagccaa tatgttccgt aaagtgcaaa
      781 ggcgggttta gtgacgttct gtttaagttt gtttgatatt ttttcttatt ccgacgtcgg
      841 ccgtaggcgt cggagattcc ggagcgtaat agggagtaga cggaacgcgg aagcgataaa
      901 aattctgcct cgtgatacgc ctatttttat aggttaatgt catgataata atggtttatt
      961 agacgtcagg tggcaccatg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     1021 tacattaaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     1081 gaaaaaggaa gagtatgagt attcaacatt tcagtgtcgc ccttattccc ttttttgcgg
     1141 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     1201 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     1261 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
     1321 gcgcggtatt ataccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt
     1381 ctcagaatga cttggttgag tactcaccag taacagaaaa gcatcttacg gatggcatga
     1441 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     1501 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     1561 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     1621 gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac
     1681 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     1741 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     1801 gtgagcgtgg atcacgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     1861 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     1921 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tacgagctcg
     1981 cttggactcc tgttgataga tccagtaatg acatcagaac tccatctgga tttgttcaga
     2041 acgctcggtt gccgccgggc gttttttatt ggtgagaatc caagcactag ggacagtaag
     2101 acgggtaagc ctgttgatga taccgctgcc ttactgggtg cattagccag tctgaatgac
     2161 ctgtcacggg ataatccgaa gtggtcagac tggaaaatca gagggcagga actgctgaac
     2221 agcaaaaagt cagatagcac cacatagcag accagccata aaacgccctg agaagcccgt
     2281 gacgggcttt tcttgtatta tgggtagttt ccttgcatga atccataaaa ggcgcctgta
     2341 gtgccattta cccacattca ctgccagagc cgtgagcgca gcgaactgaa tgtcacgaaa
     2401 aagacagcga ctcaggtgcc tgatggtcgg agacaaaagg aatattaagc gatttgcacg
     2461 agcttgcgag ggtgctactt aagcctttag ggttttaagg tctgttttgt agaggagcaa
     2521 acagcgtttg cgacatcctt ttgtaatact gcggaactga ctaaagtagt gagttataca
     2581 cagggctggg atctattctt tttatctttt tttattcttt ctttattcta taaattataa
     2641 ccacttgaat ataaacaaaa aaaacacaca aaggtctagc ggaatttaca gagggtctag
     2701 cagaatttac aagttttcca gcaaaggtct agcagaattt acagataccc acaactcaaa
     2761 ggaaaaggac atgtaattat cattgactag cccatctcaa ttggtatagt gattaaaatc
     2821 acctagacca attgagatgt atgtctgaat tagttgtttt caaagcaaat gaactagcga
     2881 ttagtcgcta tgacttaacg gagcatgaaa ccaagctaat tttatgctgt gtggcactac
     2941 tcaaccccac tattgaaaac cctacaagga aagaacggac ggtatcgttc acttataacc
     3001 aatacgctca gatgatgaac atcagtaggg aaaatgctta tggtgtatta gctaaagcaa
     3061 ccagagagct gatgacgaga actgtggaaa tcatgaatcc tttggttaaa ggctttgaga
     3121 ttttccagtg gacaaactat gccaagttct caagcgaaaa attagaatta gtttttagtg
     3181 aagagatatt gccttatctt ttccagttaa aaaaattcat aaaatataat ctggaacatg
     3241 ttaagtcttt tgaaaacaaa tactctatga ggatttatga gtggttatta aaagaactaa
     3301 cacaaaagaa aactcacaag gcaaatatag agattagcct tgatgaattt aagttcatgt
     3361 taatgcttga aaataactac catgagttta aaaggcttaa ccaatgggtt ttgaaaccaa
     3421 taagtaaaga tttaaacact tacagcaata tgaaattggt ggttgataag cgaggccgcc
     3481 cgactgatac gttgattttc caagttgaac tagatagaca aatggatctc gtaaccgaac
     3541 ttgagaacaa ccagataaaa atgaatggtg acaaaatacc aacaaccatt acatcagatt
     3601 cctacctaca taacggacta agaaaaacac tacacgatgc tttaactgca aaaattcagc
     3661 tcaccagttt tgaggcaaaa tttttgagtg acatgcaaag taagtatgat ctcaatggtt
     3721 cgttctcatg gctcacgcaa aaacaacgaa ccacactaga gaacatactg gctaaatacg
     3781 gaaggatctg aggttcttat ggctcttgta tctatcagtg aagcatcaag actaacaaac
     3841 aaaagtagaa caactgttca ccgttacata tcaaagggaa aactgtccat atgcacagat
     3901 gaaaacggtg taaaaaagat agatacatca gagcttttac gagtttttgg tgcattcaaa
     3961 gctgttcacc atgaacagat cgacaatgta acctcgagat taccgccttt gagtgagctg
     4021 ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcctgca
     4081 taacgcgaag taatcttttc ggttttaaag aaaaagggca gggtggtgac accttgccct
     4141 tctagggcgg cggatttgtc ctactcagga gagcgttcac cgacaaacaa cagataaaac
     4201 gaaaggccca gtctttcgac tgagcctttc gttttatttg atgccttaag acccactttc
     4261 acatttaagt tgtttttcta atccgcatat gatcaattca aggccgaata agaaggctgg
     4321 ctctgcacct tggtgatcaa ataattcgat agcttgtcgt aataatggcg gcatactatc
     4381 agtagtaggt gtttcccttt cttctttagc gacttgatgc tcttgatctt ccaatacgca
     4441 acctaaagta aaatgcccca ctgcgctgag tgcatataat gcattctcta gtgaaaaacc
     4501 ttgttggcat aaaaaggcta attgattttc gagagtttca tactgttttt ctgtaggccg
     4561 tgtacctaaa tgtacttttg ctccatcgcg atgacttagt aaagcacatc taaaactttt
     4621 agcgttatta cgtaaaaaat cttgccagct ttccccttct aaagggcaaa agtgagtatg
     4681 gtgcctatct aacatctcaa tggctaaggc gtcgagcaaa gcccgcttat tttttacatg
     4741 ccaatacaat gtaggctgct ctacacctag cttctgggcg agtttacggg ttgttaaacc
     4801 ttcgattccg acctcattaa gcagctctaa tgcgctgtta atcactttac ttttatctaa
     4861 acgagacatc atactacccc ctgttaaaga gtgtttctct ggctgggcta gcacaatacc
     4921 taggactgag ctagctgtca aaagcttgca tgcctgcagg tcgactctag aggatccacg
     4981 ggtaccagcg ttttgttatc aataaaaaag gccccccgat ttgggaggcc tttttagtta
     5041 gattagcgca attcaaaaag atatccttgt ccgcgtacgg tggtaatgac atcgtgcgga
     5101 tactgggcct gtattttttt ccgcagacgc cccatgagaa catcaatggt atgactttcc
     5161 cgcagttccg catccggata cagctgaagc atcagcgaat ctttgctgac cactttaccg
     5221 ttgttacgga taagcgtttc cataatggtg tattcgaacg ccgtgagttt gatgacctct
     5281 tcattgacgg ataattcccg gcgtgagaga tccacctgga acggcgggat gttgatcacc
     5341 tgggaggcca gaccgctatt acggcgcatt aacgcctgca tacgcgccat tacctcttcg
     5401 atgtggaatg gcttcgtcac gtagtcatcg gccccggagc tgagaacctc gactttatcc
     5461 tgccagcctt cgcgcgcggt taacaccaga accggcagtg aaacatcact gctgcgccag
     5521 cggcgtatta aggaaaggcc gtcttcatcc ggcagaccta aatcgacaat agcgatatcc
     5581 ggaaggtgtt cattaaggta gtaatcagct tccctggcat cttctgcggc atcgacctgg
     5641 tgacctgaat cctggagctg aaccttcagg tggtggcgta ataatgcatt atcctctaca
     5701 accagtacgc gcatcattag aaaacttcct aggcatgatt aagatgtttc agtacgaaaa
     5761 ttgctttcat tgttgatctc ctttttaagt gaacttgggc ccggtcagtg cgtcctgctg
     5821 atgtgctcag tatctctatc actgataggg atgtcaatct ctatcactga tagggagtct
     5881 tcgcgtaacg gtcgccgtta ttaaacggtt acgcgggagg aaatccttat ccgtcccgaa
     5941 aacggatata aacgcagaaa ggcccacccg aaggtgagcc agtgtgactc tagtagagag
     6001 cgttcaccga caaacaacag ataaaacgaa aggcccagtc tttcgactga gcctttcgtt
     6061 ttatttgatg cctggtctag attattagcg gccgcaacgg ctgtcgtcag tacaattatg
     6121 agtgcctggg tttatatgcg cctcatttgt acagttcatc cataccatgc gtgatgcccg
     6181 ctgcggttac gaattccagc agaaccatat gatcgcgttt ctcgttcgga tctttagaca
     6241 gaacgctttg cgtgctcaga tagtgattgt ctggcagcag aacaggacca tcaccgattg
     6301 gagtgttttg ctggtagtga tcagccagct gcacgctgcc atcctccacg ttgtggcgaa
     6361 ttttaaaatt cgctttaatg ccattttttt gtttatcggc ggtgatgtaa acattgtggc
     6421 t
//
