LOCUS       sequence-335739-        6237 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes PslyB-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 6237)
  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 6237)
  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: pAM007
FEATURES             Location/Qualifiers
     source          1..6237
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(19..123)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     188..206
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     RBS             474..485
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             492..1205
                     /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"
     terminator      1284..1355
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1371..1398
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     promoter        join(1475..1493,1494..1499,1500..1516,1517..1522,
                     1523..1548)
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    1475..1493
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1500..1518
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        2410..2444
                     /label=BBa_J23104
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.72"
     CDS             2479..3105
                     /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     complement(2555..2574)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     terminator      3111..3197
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(3281..3298)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             complement(3560..4510)
                     /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      4558..4780
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(5272..5366)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(5396..6184)
                     /codon_start=1
                     /gene="bla(M)"
                     /product="beta-lactamase lacking the signal sequence"
                     /label=bla(M)
                     /note="allows cytosolic expression of beta-lactamase"
                     /translation="PETLVKVKDAEDQLGARVGYIELDLNSGKILESFRPEERFPMMST
                     FKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYSPVTEKHLTDGMTVRELCSAAITMS
                     DNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRWEPELNEAIPNDERDTTMPVAMATT
                     LRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSALPAGWFIADKSGAGERGSRGIIAA
                     LGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGASLIKHW"
     primer_bind     6019..6038
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
       61 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
      121 gcgcacattt cccatggtgc cacctgacgt ctaagaaacc attattatca tgacattaac
      181 ctataaaaat aggcgtatca cgaggcagaa tttttatcgc ttccgcgttc cgtctactcc
      241 ctattacgct ccggaatctc cgacgcctac ggccgacgtc ggaccgtgtc gtccacggat
      301 atcaatgacc tatcttttct tgcttccgac ttcgtttaag attggttaat taacttctga
      361 ttatgatttt cacattcttt cggcagaaga tatgacctta actgcataaa tgccatataa
      421 tttagctata gggatgctga aatagcgttt cgtgaagaac aggagatggg taaaaagagg
      481 agaaatacta gatgcgtaaa ggcgaagagc tgttcactgg tgtcgtccct attctggtgg
      541 aactggatgg tgatgtcaac ggtcataagt tttccgtgcg tggcgagggt gaaggtgacg
      601 caactaatgg taaactgacg ctgaagttca tctgtactac tggtaaactg ccggtacctt
      661 ggccgactct ggtaacgacg ctgacttatg gtgttcagtg ctttgctcgt tatccggacc
      721 atatgaagca gcatgacttc ttcaagtccg ccatgccgga aggctatgtg caggaacgca
      781 cgatttcctt taaggatgac ggcacgtaca aaacgcgtgc ggaagtgaaa tttgaaggcg
      841 ataccctggt aaaccgcatt gagctgaaag gcattgactt taaagaagac ggcaatatcc
      901 tgggccataa gctggaatac aattttaaca gccacaatgt ttacatcacc gccgataaac
      961 aaaaaaatgg cattaaagcg aattttaaaa ttcgccacaa cgtggaggat ggcagcgtgc
     1021 agctggctga tcactaccag caaaacactc caatcggtga tggtcctgtt ctgctgccag
     1081 acaatcacta tctgagcacg caaagcgttc tgtctaaaga tccgaacgag aaacgcgatc
     1141 atatggttct gctggaattc gtaaccgcag cgggcatcac gcatggtatg gatgaactgt
     1201 acaaatgagg cgcatataaa cccaggcact cataattgta ctgacgacag ccgttgcggc
     1261 cgctaataat ctagaccagg catcaaataa aacgaaaggc tcagtcgaaa gactgggcct
     1321 ttcgttttat ctgttgtttg tcggtgaacg ctctctacta gagtcacact ggctcacctt
     1381 cgggtgggcc tttctgcgtt tatatccgtt ttcgggacgg ataaggattt cctcccgcgt
     1441 aaccgtttaa taacggcgac cgttacgcga agactcccta tcagtgatag agattgacat
     1501 ccctatcagt gatagagata ctgagcacat cagcaggacg cactgaccgg gcccaagttc
     1561 acttaaaaag gagatcaaca atgaaagcaa ttttcgtact gaaacatctt aatcatgcct
     1621 aggaagtttt ctaatgatgc gcgtactggt tgtagaggat aatgcattat tacgccacca
     1681 cctgaaggtt cagctccagg attcaggtca ccaggtcgat gccgcagaag atgccaggga
     1741 agctgattac taccttaatg aacaccttcc ggatatcgct attgtcgatt taggtctgcc
     1801 ggatgaagac ggcctttcct taatacgccg ctggcgcagc agtgatgttt cactgccggt
     1861 tctggtgtta accgcgcgcg aaggctggca ggataaagtc gaggttctca gctccggggc
     1921 cgatgactac gtgacgaagc cattccacat cgaagaggta atggcgcgta tgcaggcgtt
     1981 aatgcgccgt aatagcggtc tggcctccca ggtgatcaac atcccgccgt tccaggtgga
     2041 tctctcacgc cgggaattat ccgtcaatga agaggtcatc aaactcacgg cgttcgaata
     2101 caccattatg gaaacgctta tccgtaacaa cggtaaagtg gtcagcaaag attcgctgat
     2161 gcttcagctg tatccggatg cggaactgcg ggaaagtcat accattgatg ttctcatggg
     2221 gcgtctgcgg aaaaaaatac aggcccagta tccgcacgat gtcattacca ccgtacgcgg
     2281 acaaggatat ctttttgaat tgcgctaatc taactaaaaa ggcctcccaa atcggggggc
     2341 cttttttatt gataacaaaa cgctggtacc cgtggatcct ctagagtcga cctgcaggca
     2401 tgcaagcttt tgacagctag ctcagtccta ggtattgtgc tagcccagcc agagaaacac
     2461 tctttaacag ggggtagtat gatgtctcgt ttagataaaa gtaaagtgat taacagcgca
     2521 ttagagctgc ttaatgaggt cggaatcgaa ggtttaacaa cccgtaaact cgcccagaag
     2581 ctaggtgtag agcagcctac attgtattgg catgtaaaaa ataagcgggc tttgctcgac
     2641 gccttagcca ttgagatgtt agataggcac catactcact tttgcccttt agaaggggaa
     2701 agctggcaag attttttacg taataacgct aaaagtttta gatgtgcttt actaagtcat
     2761 cgcgatggag caaaagtaca tttaggtaca cggcctacag aaaaacagta tgaaactctc
     2821 gaaaatcaat tagccttttt atgccaacaa ggtttttcac tagagaatgc attatatgca
     2881 ctcagcgcag tggggcattt tactttaggt tgcgtattgg aagatcaaga gcatcaagtc
     2941 gctaaagaag aaagggaaac acctactact gatagtatgc cgccattatt acgacaagct
     3001 atcgaattat ttgatcacca aggtgcagag ccagccttct tattcggcct tgaattgatc
     3061 atatgcggat tagaaaaaca acttaaatgt gaaagtgggt cttaaggcat caaataaaac
     3121 gaaaggctca gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg gtgaacgctc
     3181 tcctgagtag gacaaatccg ccgccctaga agggcaaggt gtcaccaccc tgcccttttt
     3241 ctttaaaacc gaaaagatta cttcgcgtta tgcaggcttc ctcgctcact gactcgctgc
     3301 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atctcgaggt
     3361 tacattgtcg atctgttcat ggtgaacagc tttaaatgca ccaaaaactc gtaaaagctc
     3421 tgatgtatct atctttttta caccgttttc atctgtgcat atggacagtt ttccctttga
     3481 tatctaacgg tgaacagttg ttctactttt gtttgttagt cttgatgctt cactgataga
     3541 tacaagagcc ataagaacct cagatccttc cgtatttagc cagtatgttc tctagtgtgg
     3601 ttcgttgttt ttgcgtgagc catgagaacg aaccattgag atcatgctta ctttgcatgt
     3661 cactcaaaaa ttttgcctca aaactggtga gctgaatttt tgcagttaaa gcatcgtgta
     3721 gtgtttttct tagtccgtta cgtaggtagg aatctgatgt aatggttgtt ggtattttgt
     3781 caccattcat ttttatctgg ttgttctcaa gttcggttac gagatccatt tgtctatcta
     3841 gttcaacttg gaaaatcaac gtatcagtcg ggcggcctcg cttatcaacc accaatttca
     3901 tattgctgta agtgtttaaa tctttactta ttggtttcaa aacccattgg ttaagccttt
     3961 taaactcatg gtagttattt tcaagcatta acatgaactt aaattcatca aggctaatct
     4021 ctatatttgc cttgtgagtt ttcttttgtg ttagttcttt taataaccac tcataaatcc
     4081 tcatagagta tttgttttca aaagacttaa catgttccag attatatttt atgaattttt
     4141 ttaactggaa aagataaggc aatatctctt cactaaaaac taattctaat ttttcgcttg
     4201 agaacttggc atagtttgtc cactggaaaa tctcaaagcc tttaaccaaa ggattcctga
     4261 tttccacagt tctcgtcatc agctctctgg ttgctttagc taatacacca taagcatttt
     4321 ccctactgat gttcatcatc tgagcgtatt ggttataagt gaacgatacc gtccgttctt
     4381 tccttgtagg gttttcaatc gtggggttga gtagtgccac acagcataaa attagcttgg
     4441 tttcatgctc cgttaagtca tagcgactaa tcgctagttc atttgctttg aaaacaacta
     4501 attcagacat acatctcaat tggtctaggt gattttaatc actataccaa ttgagatggg
     4561 ctagtcaatg ataattacat gtccttttcc tttgagttgt gggtatctgt aaattctgct
     4621 agacctttgc tggaaaactt gtaaattctg ctagaccctc tgtaaattcc gctagacctt
     4681 tgtgtgtttt ttttgtttat attcaagtgg ttataattta tagaataaag aaagaataaa
     4741 aaaagataaa aagaatagat cccagccctg tgtataactc actactttag tcagttccgc
     4801 agtattacaa aaggatgtcg caaacgctgt ttgctcctct acaaaacaga ccttaaaacc
     4861 ctaaaggctt aagtagcacc ctcgcaagct cgggcaaatc gctgaatatt ccttttgtct
     4921 ccgaccatca ggcacctgag tcgctgtctt tttcgtgaca ttcagttcgc tgcgctcacg
     4981 gctctggcag tgaatggggg taaatggcac tacaggcgcc ttttatggat tcatgcaagg
     5041 aaactaccca taatacaaga aaagcccgtc acgggcttct cagggcgttt tatggcgggt
     5101 ctgctatgtg gtgctatctg actttttgct gttcagcagt tcctgccctc tgattttcca
     5161 gtctgaccac ttcggattat cccgtgacag gtcattcaga ctggctaatg cacccagtaa
     5221 ggcagcggta tcatcaacag gcttacccgt cttactgtcc ctagtgcttg gattctcacc
     5281 aataaaaaac gcccggcggc aaccgagcgt tctgaacaaa tccagatgga gttctgaggt
     5341 cattactgga tctatcaaca ggagtccaag cgagctcgta aacttggtct gacagttacc
     5401 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     5461 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     5521 ctgcaatgat accgcgtgat ccacgctcac cggctccaga tttatcagca ataaaccagc
     5581 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     5641 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     5701 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     5761 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     5821 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     5881 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     5941 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     6001 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     6061 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     6121 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     6181 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacac
//
