LOCUS       sequence-335938-        6697 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes PpagC-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 6697)
  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 6697)
  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: pAM006
FEATURES             Location/Qualifiers
     source          1..6697
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             463..474
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     primer_bind     complement(1201..1219)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1284..1388
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1389..1457,1458..2249)
                     /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(1607..1626)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      2279..2373
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     rep_origin      2865..3087
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     CDS             3135..4085
                     /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     4347..4364
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(4448..4534)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(4540..5166)
                     /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     5071..5090
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        complement(5201..5235)
                     /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(6097..6122,6123..6128,6129..6145,6146..6151,
                     6152..6170))
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    6127..6145
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    6152..6170
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     terminator      complement(6247..6274)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      complement(6290..6361)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(join(6440..6697,1..456))
                     /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 gatgtaaaca ttgtggctgt taaaattgta ttccagctta tggcccagga tattgccgtc
       61 ttctttaaag tcaatgcctt tcagctcaat gcggtttacc agggtatcgc cttcaaattt
      121 cacttccgca cgcgttttgt acgtgccgtc atccttaaag gaaatcgtgc gttcctgcac
      181 atagccttcc ggcatggcgg acttgaagaa gtcatgctgc ttcatatggt ccggataacg
      241 agcaaagcac tgaacaccat aagtcagcgt cgttaccaga gtcggccaag gtaccggcag
      301 tttaccagta gtacagatga acttcagcgt cagtttacca ttagttgcgt caccttcacc
      361 ctcgccacgc acggaaaact tatgaccgtt gacatcacca tccagttcca ccagaatagg
      421 gacgacacca gtgaacagct cttcgccttt acgcatctag tatttctcct ctttaacaac
      481 tccttaatac tacttattat ttacggtgtg tttaaacacc tgcagtaccg atccggcatt
      541 cagttatcgc cactatgccg aatcgacaaa accacgaata attcaccgct atcgctcctg
      601 atgtgtttac ttcctgaaag atatttttac taccgaagca ctctatcgct catttaggta
      661 accggttcta caatgtcatc taacttttat agatttgaat gctaattttt ctcacgcata
      721 tatatttaac agaaaccata aagtgtttag ccactataga acaacaaatc acccatgcaa
      781 cattttgata tttaaagaga aaatctcaca accacattaa gaaacttgac accgttcggc
      841 taaaaacatg tcattaagca aactcgccat ataatcagaa catatcgcat tgtgcttcac
      901 agtcctcacg tgacgctcca tccgcaatac ggttatatgc catcgcaggc gctgtaatca
      961 tattcacgat gatgcttagc acgctttatt cccgctccga tttaatcttt taatatatct
     1021 atcagttaca acatttcttg ttatattata agaatagaat caacaccaca attccaacat
     1081 aaatatcacc tgtgtttaga gagaatttac attccaaaaa aattccgacg tcggccgtag
     1141 gcgtcggaga ttccggagcg taatagggag tagacggaac gcggaagcga taaaaattct
     1201 gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt tattagacgt
     1261 caggtggcac catgggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
     1321 aaaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
     1381 ggaagagtat gagtattcaa catttcagtg tcgcccttat tccctttttt gcggcatttt
     1441 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
     1501 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
     1561 ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
     1621 tattataccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
     1681 atgacttggt tgagtactca ccagtaacag aaaagcatct tacggatggc atgacagtaa
     1741 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
     1801 caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
     1861 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
     1921 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
     1981 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
     2041 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
     2101 gtggatcacg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
     2161 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
     2221 taggtgcctc actgattaag cattggtaac tgtcagacca agtttacgag ctcgcttgga
     2281 ctcctgttga tagatccagt aatgacatca gaactccatc tggatttgtt cagaacgctc
     2341 ggttgccgcc gggcgttttt tattggtgag aatccaagca ctagggacag taagacgggt
     2401 aagcctgttg atgataccgc tgccttactg ggtgcattag ccagtctgaa tgacctgtca
     2461 cgggataatc cgaagtggtc agactggaaa atcagagggc aggaactgct gaacagcaaa
     2521 aagtcagata gcaccacata gcagaccagc cataaaacgc cctgagaagc ccgtgacggg
     2581 cttttcttgt attatgggta gtttccttgc atgaatccat aaaaggcgcc tgtagtgcca
     2641 tttacccaca ttcactgcca gagccgtgag cgcagcgaac tgaatgtcac gaaaaagaca
     2701 gcgactcagg tgcctgatgg tcggagacaa aaggaatatt aagcgatttg cacgagcttg
     2761 cgagggtgct acttaagcct ttagggtttt aaggtctgtt ttgtagagga gcaaacagcg
     2821 tttgcgacat ccttttgtaa tactgcggaa ctgactaaag tagtgagtta tacacagggc
     2881 tgggatctat tctttttatc tttttttatt ctttctttat tctataaatt ataaccactt
     2941 gaatataaac aaaaaaaaca cacaaaggtc tagcggaatt tacagagggt ctagcagaat
     3001 ttacaagttt tccagcaaag gtctagcaga atttacagat acccacaact caaaggaaaa
     3061 ggacatgtaa ttatcattga ctagcccatc tcaattggta tagtgattaa aatcacctag
     3121 accaattgag atgtatgtct gaattagttg ttttcaaagc aaatgaacta gcgattagtc
     3181 gctatgactt aacggagcat gaaaccaagc taattttatg ctgtgtggca ctactcaacc
     3241 ccactattga aaaccctaca aggaaagaac ggacggtatc gttcacttat aaccaatacg
     3301 ctcagatgat gaacatcagt agggaaaatg cttatggtgt attagctaaa gcaaccagag
     3361 agctgatgac gagaactgtg gaaatcatga atcctttggt taaaggcttt gagattttcc
     3421 agtggacaaa ctatgccaag ttctcaagcg aaaaattaga attagttttt agtgaagaga
     3481 tattgcctta tcttttccag ttaaaaaaat tcataaaata taatctggaa catgttaagt
     3541 cttttgaaaa caaatactct atgaggattt atgagtggtt attaaaagaa ctaacacaaa
     3601 agaaaactca caaggcaaat atagagatta gccttgatga atttaagttc atgttaatgc
     3661 ttgaaaataa ctaccatgag tttaaaaggc ttaaccaatg ggttttgaaa ccaataagta
     3721 aagatttaaa cacttacagc aatatgaaat tggtggttga taagcgaggc cgcccgactg
     3781 atacgttgat tttccaagtt gaactagata gacaaatgga tctcgtaacc gaacttgaga
     3841 acaaccagat aaaaatgaat ggtgacaaaa taccaacaac cattacatca gattcctacc
     3901 tacataacgg actaagaaaa acactacacg atgctttaac tgcaaaaatt cagctcacca
     3961 gttttgaggc aaaatttttg agtgacatgc aaagtaagta tgatctcaat ggttcgttct
     4021 catggctcac gcaaaaacaa cgaaccacac tagagaacat actggctaaa tacggaagga
     4081 tctgaggttc ttatggctct tgtatctatc agtgaagcat caagactaac aaacaaaagt
     4141 agaacaactg ttcaccgtta catatcaaag ggaaaactgt ccatatgcac agatgaaaac
     4201 ggtgtaaaaa agatagatac atcagagctt ttacgagttt ttggtgcatt caaagctgtt
     4261 caccatgaac agatcgacaa tgtaacctcg agattaccgc ctttgagtga gctgataccg
     4321 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcc tgcataacgc
     4381 gaagtaatct tttcggtttt aaagaaaaag ggcagggtgg tgacaccttg cccttctagg
     4441 gcggcggatt tgtcctactc aggagagcgt tcaccgacaa acaacagata aaacgaaagg
     4501 cccagtcttt cgactgagcc tttcgtttta tttgatgcct taagacccac tttcacattt
     4561 aagttgtttt tctaatccgc atatgatcaa ttcaaggccg aataagaagg ctggctctgc
     4621 accttggtga tcaaataatt cgatagcttg tcgtaataat ggcggcatac tatcagtagt
     4681 aggtgtttcc ctttcttctt tagcgacttg atgctcttga tcttccaata cgcaacctaa
     4741 agtaaaatgc cccactgcgc tgagtgcata taatgcattc tctagtgaaa aaccttgttg
     4801 gcataaaaag gctaattgat tttcgagagt ttcatactgt ttttctgtag gccgtgtacc
     4861 taaatgtact tttgctccat cgcgatgact tagtaaagca catctaaaac ttttagcgtt
     4921 attacgtaaa aaatcttgcc agctttcccc ttctaaaggg caaaagtgag tatggtgcct
     4981 atctaacatc tcaatggcta aggcgtcgag caaagcccgc ttatttttta catgccaata
     5041 caatgtaggc tgctctacac ctagcttctg ggcgagttta cgggttgtta aaccttcgat
     5101 tccgacctca ttaagcagct ctaatgcgct gttaatcact ttacttttat ctaaacgaga
     5161 catcatacta ccccctgtta aagagtgttt ctctggctgg gctagcacaa tacctaggac
     5221 tgagctagct gtcaaaagct tgcatgcctg caggtcgact ctagaggatc cacgggtacc
     5281 agcgttttgt tatcaataaa aaaggccccc cgatttggga ggccttttta gttagattag
     5341 cgcaattcaa aaagatatcc ttgtccgcgt acggtggtaa tgacatcgtg cggatactgg
     5401 gcctgtattt ttttccgcag acgccccatg agaacatcaa tggtatgact ttcccgcagt
     5461 tccgcatccg gatacagctg aagcatcagc gaatctttgc tgaccacttt accgttgtta
     5521 cggataagcg tttccataat ggtgtattcg aacgccgtga gtttgatgac ctcttcattg
     5581 acggataatt cccggcgtga gagatccacc tggaacggcg ggatgttgat cacctgggag
     5641 gccagaccgc tattacggcg cattaacgcc tgcatacgcg ccattacctc ttcgatgtgg
     5701 aatggcttcg tcacgtagtc atcggccccg gagctgagaa cctcgacttt atcctgccag
     5761 ccttcgcgcg cggttaacac cagaaccggc agtgaaacat cactgctgcg ccagcggcgt
     5821 attaaggaaa ggccgtcttc atccggcaga cctaaatcga caatagcgat atccggaagg
     5881 tgttcattaa ggtagtaatc agcttccctg gcatcttctg cggcatcgac ctggtgacct
     5941 gaatcctgga gctgaacctt caggtggtgg cgtaataatg cattatcctc tacaaccagt
     6001 acgcgcatca ttagaaaact tcctaggcat gattaagatg tttcagtacg aaaattgctt
     6061 tcattgttga tctccttttt aagtgaactt gggcccggtc agtgcgtcct gctgatgtgc
     6121 tcagtatctc tatcactgat agggatgtca atctctatca ctgataggga gtcttcgcgt
     6181 aacggtcgcc gttattaaac ggttacgcgg gaggaaatcc ttatccgtcc cgaaaacgga
     6241 tataaacgca gaaaggccca cccgaaggtg agccagtgtg actctagtag agagcgttca
     6301 ccgacaaaca acagataaaa cgaaaggccc agtctttcga ctgagccttt cgttttattt
     6361 gatgcctggt ctagattatt agcggccgca acggctgtcg tcagtacaat tatgagtgcc
     6421 tgggtttata tgcgcctcat ttgtacagtt catccatacc atgcgtgatg cccgctgcgg
     6481 ttacgaattc cagcagaacc atatgatcgc gtttctcgtt cggatcttta gacagaacgc
     6541 tttgcgtgct cagatagtga ttgtctggca gcagaacagg accatcaccg attggagtgt
     6601 tttgctggta gtgatcagcc agctgcacgc tgccatcctc cacgttgtgg cgaattttaa
     6661 aattcgcttt aatgccattt ttttgtttat cggcggt
//
