LOCUS       sequence-436632-        7737 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To track the natural CRP promoter's transcriptional dynamics.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7737)
  AUTHORS   Dash S, Jagadeesan R, Baptista ISC, Chauhan V, Kandavalli V, 
            Oliveira SMD, Ribeiro AS
  TITLE     A library of reporters of the global regulators of gene expression 
            in Escherichia coli.
  JOURNAL   mSystems. 2024 Jun 18;9(6):e0006524. doi: 10.1128/msystems.00065-24.
            Epub 2024 Apr 30.
  PUBMED    38687030
REFERENCE   2  (bases 1 to 7737)
  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: "mSystems.";
            date: "2024-06-18"; volume: "9(6):e0006524. doi"; pages: " 
            10.1128/msystems.00065-24. Epub 2024 Apr 30"
COMMENT     Sequence Label: CRP-GFP
FEATURES             Location/Qualifiers
     source          1..7737
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(391..1566)
                     /codon_start=1
                     /gene="sopA"
                     /product="partitioning protein for the bacterial F plasmid"
                     /label=sopA
                     /translation="MFRMKLMETLNQCINAGHEMTKAIAIAQFNDDSPEARKITRRWRI
                     GEAADLVGVSSQAIRDAEKAGRLPHPDMEIRGRVEQRVGYTIEQINHMRDVFGTRLRRA
                     EDVFPPVIGVAAHKGGVYKTSVSVHLAQDLALKGLRVLLVEGNDPQGTASMYHGWVPDL
                     HIHAEDTLLPFYLGEKDDVTYAIKPTCWPGLDIIPSCLALHRIETELMGKFDEGKLPTD
                     PHLMLRLAIETVAHDYDVIVIDSAPNLGIGTINVVCAADVLIVPTPAELFDYTSALQFF
                     DMLRDLLKNVDLKGFEPDVRILLTKYSNSNGSQSPWMEEQIRDAWGSMVLKNVVRETDE
                     VGKGQIRMRTVFEQAIDQRSSTGAWRNALSIWEPVCNEIFDRLIKPRWEIR"
     misc_feature    1892..2142
                     /gene="incC"
                     /label=incC
                     /note="incompatibility region of the bacterial F plasmid"
     CDS             complement(2145..2900)
                     /codon_start=1
                     /gene="repE"
                     /product="replication initiation protein for the bacterial 
                     F plasmid"
                     /label=repE
                     /translation="MAETAVINHKKRKNSPRIVQSNDLTEAAYSLSRDQKRMLYLFVDQ
                     IRKSDGTLQEHDGICEIHVAKYAEIFGLTSAEASKDIRQALKSFAGKEVVFYRPEEDAG
                     DEKGYESFPWFIKRAHSPSRGLYSVHINPYLIPFFIGLQNRFTQFRLSETKEITNPYAM
                     RLYESLCQYRKPDGSGIVSLKIDWIIERYQLPQSYQRMPDFRRRFLQVCVNEINSRTPM
                     RLSYIEKKKGRQTTHIVFSFRDITSMTTG"
     rep_origin      2991..3210
                     /label=ori2
                     /note="secondary origin of replication for the bacterial F 
                     plasmid; also known as oriS"
     rep_origin      3278..3809
                     /label=oriV
                     /note="incP origin of replication"
     rep_origin      3286..3900
                     /label=oriV
                     /note="origin of replication for the bacterial F plasmid"
     promoter        4758..4860
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             4861..5520
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(4927..4946)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      complement(5655..5749)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(5812..6525)
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(5852..5872)
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     6467..6495
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     RBS             6531..6539
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     terminator      6913..6999
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(join(7157..7737,1..391))
                     /codon_start=1
                     /gene="sopB"
                     /product="partitioning protein for the bacterial F plasmid"
                     /label=sopB
                     /translation="MKRAPVIPKHTLNTQPVEDTSLSTPAAPMVDSLIARVGVMARGNA
                     ITLPVCGRDVKFTLEVLRGDSVEKTSRVWSGNERDQELLTEDALDDLIPSFLLTGQQTP
                     AFGRRVSGVIEIADGSRRRKAAALTESDYRVLVGELDDEQMAALSRLGNDYRPTSAYER
                     GQRYASRLQNEFAGNISALADAENISRKIITRCINTAKLPKSVVALFSHPGELSARSGD
                     ALQKAFTDKEELLKQQASNLHEQKKAGVIFEAEEVITLLTSVLKTSSASRTSLSSRHQF
                     APGATVLYKGDKMVLNLDRSRVPTECIEKIEAILKELEKPAP"
ORIGIN
        1 tttcggtaag tgcagcagct ttacgacggc gactcccatc ggcaatttct atgacaccag
       61 atactcttcg accgaacgcc ggtgtctgtt gaccagtcag tagaaaagaa gggatgagat
      121 catccagtgc gtcctcagta agcagctcct ggtcacgttc attacctgac catacccgag
      181 aggtcttctc aacactatca ccccggagca cttcaagagt aaacttcaca tcccgaccac
      241 atacaggcaa agtaatggca ttaccgcgag ccattactcc tacgcgcgca attaacgaat
      301 ccaccatcgg ggcagctggt gtcgataacg aagtatcttc aaccggttga gtattgagcg
      361 tatgttttgg aataacaggc gcacgcttca ttatctaatc tcccagcgtg gtttaatcag
      421 acgatcgaaa atttcattgc agacaggttc ccaaatagaa agagcatttc tccaggcacc
      481 agttgaagag cgttgatcaa tggcctgttc aaaaacagtt ctcatccgga tctgaccttt
      541 accaacttca tccgtttcac gtacaacatt ttttagaacc atgcttcccc aggcatcccg
      601 aatttgctcc tccatccacg gggactgaga gccattacta ttgctgtatt tggtaagcaa
      661 aatacgtaca tcaggctcga accctttaag atcaacgttc ttgagcagat cacgaagcat
      721 atcgaaaaac tgcagtgcgg aggtgtagtc aaacaactca gcaggcgtgg gaacaatcag
      781 cacatcagca gcacatacga cattaatcgt gccgataccc aggttaggcg cgctgtcaat
      841 aactatgaca tcatagtcat gagcaacagt ttcaatggcc agtcggagca tcaggtgtgg
      901 atcggtgggc agtttacctt catcaaattt gcccattaac tcagtttcaa tacggtgcag
      961 agccagacag gaaggaataa tgtcaagccc cggccagcaa gtgggcttta ttgcataagt
     1021 gacatcgtcc ttttccccaa gatagaaagg caggagagtg tcttctgcat gaatatgaag
     1081 atctggtacc catccgtgat acattgaggc tgttccctgg gggtcgttac cttccacgag
     1141 caaaacacgt agccccttca gagccagatc ctgagcaaga tgaacagaaa ctgaggtttt
     1201 gtaaacgcca cctttatggg cagcaacccc gatcaccggt ggaaatacgt cttcagcacg
     1261 tcgcaatcgc gtaccaaaca catcacgcat atgattaatt tgttcaattg tataaccaac
     1321 acgttgctca acccgtcctc gaatttccat atccgggtgc ggtagtcgcc ctgctttctc
     1381 ggcatctctg atagcctgag aagaaacccc aactaaatcc gctgcttcac ctattctcca
     1441 gcgccgggtt attttcctcg cttccgggct gtcatcatta aactgtgcaa tggcgatagc
     1501 cttcgtcatt tcatgaccag cgtttatgca ctggttaagt gtttccatga gtttcattct
     1561 gaacatcctt taatcattgc tttgcgtttt tttattaaat cttgcaattt actgcaaagc
     1621 aacaacaaaa tcgcaaagtc atcaaaaaac cgcaaagttg tttaaaataa gagcaacact
     1681 acaaaaggag ataagaagag cacatacctc agtcacttat tatcactagc gctcgccgca
     1741 gccgtgtaac cgagcatagc gagcgaactg gcgaggaagc aaagaagaac tgttctgtca
     1801 gatagctctt acgctcagcg caagaagaaa tatccaccgt gggaaaaact ccaggtagag
     1861 gtacacacgc ggatagccaa ttcagagtaa taaactgtga taatcaaccc tcatcaatga
     1921 tgacgaacta acccccgata tcaagtcaca tgacgaaggg aaagagaagg aaatcaactg
     1981 tgacaaactg ccctcaaatt tggcttcctt aaaaattaca gttcaaaaag tatgagaaaa
     2041 tccatgcagg ctgaaggaaa cagcaaaact gtgacaaatt accctcagta ggtcagaaca
     2101 aatgtgacga accaccctca aatctgtgac agataaccct cagactatcc tgtcgtcatg
     2161 gaagtgatat cgcggaagga aaatacgata tgagtcgtct ggcggccttt ctttttctca
     2221 atgtatgaga ggcgcattgg agttctgctg ttgatctcat taacacagac ctgcaggaag
     2281 cggcggcgga agtcaggcat acgctggtaa ctttgaggca gctggtaacg ctctatgatc
     2341 cagtcgattt tcagagagac gatgcctgag ccatccggct tacgatactg acacagggat
     2401 tcgtataaac gcatggcata cggattggtg atttcttttg tttcactaag ccgaaactgc
     2461 gtaaaccggt tctgtaaccc gataaagaag ggaatgagat atgggttgat atgtacactg
     2521 taaagccctc tggatggact gtgcgcacgt ttgataaacc aaggaaaaga ttcatagcct
     2581 ttttcatcgc cggcatcctc ttcagggcga taaaaaacca cttccttccc cgcgaaactc
     2641 ttcaatgcct gccgtatatc cttactggct tccgcagagg tcaatccgaa tatttcagca
     2701 tatttagcaa catggatctc gcagataccg tcatgttcct gtagggtgcc atcagatttt
     2761 ctgatctggt caacgaacag atacagcata cgtttttgat cccgggagag actatatgcc
     2821 gcctcagtga ggtcgtttga ctggacgatt cgcgggctat ttttacgttt cttgtgattg
     2881 ataaccgctg tttccgccat gacagatcca tgtgaagtgt gacaagtttt tagattgtca
     2941 cactaaataa aaaagagtca ataagcaggg ataactttgt gaaaaaacag cttcttctga
     3001 gggcaatttg tcacagggtt aagggcaatt tgtcacagac aggactgtca tttgagggtg
     3061 atttgtcaca ctgaaagggc aatttgtcac aacaccttct ctagaaccag catggataaa
     3121 ggcctacaag gcgctctaaa aaagaagatc taaaaactat aaaaaaaata attataaaaa
     3181 tatccccgtg gataagtgga taaccccaag ggaagttttt tcaggcatcg tgtgtaagca
     3241 gaatatataa gtgctgttcc ctggtgcttc ctcgctcact cgaccgggag ggttcgagaa
     3301 gggggggcac cccccttcgg cgtgcgcggt cacgcgcaca gggcgcagcc ctggttaaaa
     3361 acaaggttta taaatattgg tttaaaagca ggttaaaaga caggttagcg gtggccgaaa
     3421 aacgggcgga aacccttgca aatgctggat tttctgcctg tggacagccc ctcaaatgtc
     3481 aataggtgcg cccctcatct gtcagcactc tgcccctcaa gtgtcaagga tcgcgcccct
     3541 catctgtcag tagtcgcgcc cctcaagtgt caataccgca gggcacttat ccccaggctt
     3601 gtccacatca tctgtgggaa actcgcgtaa aatcaggcgt tttcgccgat ttgcgaggct
     3661 ggccagctcc acgtcgccgg ccgaaatcga gcctgcccct catctgtcaa cgccgcgccg
     3721 ggtgagtcgg cccctcaagt gtcaacgtcc gcccctcatc tgtcagtgag ggccaagttt
     3781 tccgcgaggt atccacaacg ccggcggccg gccgcggtgt ctcgcacacg gcttcgacgg
     3841 cgtttctggc gcgtttgcag ggccatagac ggccgccagc ccagcggcga gggcaaccag
     3901 ccgagggctt cgccctgtcg ctcgactgcg gcgagcacta ctggctgtaa aaggacagac
     3961 cacatcatgg ttctgtgttc attaggttgt tctgtccatt gctgacataa tccgctccac
     4021 ttcaacgtaa caccgcacga agatttctat tgttcctgaa ggcatattca aatcgttttc
     4081 gttaccgctt gctggcatca tgacagaaca ctacttccta taaacgctac acaggctcct
     4141 gagattaata atgcggatct ctacgataat gggagatttt cccgactgtt tcgttcgctt
     4201 ctcagtggat aacagccagc ttctctgttt aacagacaaa aacagcatat ccactcagtt
     4261 ccacatttcc atataaaggc caaggcattt attctcagga taattgtttc agcatcgcaa
     4321 ccgcatcaga ctccggcatc gcaaactgca cccggtgccg ggcagccaca tccagcgcaa
     4381 aaaccttcgt gtagacttcc gttgaactga tggacttatg tcccatcagg ctttgcagaa
     4441 ctttcagcgg tataccggca tacagcatgt gcatcgcata ggaatggcgg aacgtatgtg
     4501 gtgtgaccgg aacagagaac gtcacaccgt cagcagcagc ggcggcaacc gcctccccaa
     4561 tccaggtcct gaccgttctg tccgtcactt cccagatccg cgctttctct gtccttcctg
     4621 tgcgacggtt acgccgctcc atgagcttat cgcgaataaa tacctgtgac ggaagatcac
     4681 ttcgcagaat aaataaatcc tggtgtccct gttgataccg ggaagccctg ggccaacttt
     4741 tggcgaaaat gagacgttga tcggcacgta agaggttcca actttcacca taatgaaata
     4801 agatcactac cgggcgtatt ttttgagtta tcgagatttt caggagctaa ggaagctaaa
     4861 atggagaaaa aaatcactgg atataccacc gttgatatat cccaatggca tcgtaaagaa
     4921 cattttgagg catttcagtc agttgctcaa tgtacctata accagaccgt tcagctggat
     4981 attacggcct ttttaaagac cgtaaagaaa aataagcaca agttttatcc ggcctttatt
     5041 cacattcttg cccgcctgat gaatgctcat ccggaatttc gtatggcaat gaaagacggt
     5101 gagctggtga tatgggatag tgttcaccct tgttacaccg ttttccatga gcaaactgaa
     5161 acgttttcat cgctctggag tgaataccac gacgatttcc ggcagtttct acacatatat
     5221 tcgcaagatg tggcgtgtta cggtgaaaac ctggcctatt tccctaaagg gtttattgag
     5281 aatatgtttt tcgtctcagc caatccctgg gtgagtttca ccagttttga tttaaacgtg
     5341 gccaatatgg acaacttctt cgcccccgtt ttcacgatgg gcaaatatta tacgcaaggc
     5401 gacaaggtgc tgatgccgct ggcgattcag gttcatcatg ccgtttgtga tggcttccat
     5461 gtcggcagaa tgcttaatga attacaacag tactgcgatg agtggcaggg cggggcgtaa
     5521 tttttttaag gcagttattg gtgcccttaa acgcctggtt gctacgcctg aataagtgat
     5581 aataagcgga tgaatggcag aaattcgatg ataagctgtc aaacatgaga attggtctcg
     5641 acgagtatgc ttggattctc accaataaaa aacgcccggc ggcaaccgag cgttctgaac
     5701 aaatccagat ggagttctga ggtcattact ggatctatca acaggagtcc aagcgacgtc
     5761 tgtgcaagta ctactgttct gcagtcactt gaattcgata cccagctgtt atttgtatag
     5821 ttcatccatg ccatgtgtaa tcccagcagc ggttacaaac tcaagaagga ccatgtggtc
     5881 tctcttttcg ttgggatctt tcgaaagggc agattgtgtg gacaggtaat ggttgtctgg
     5941 taaaaggaca gggccatcgc caatcggagt attttgttga taatggtctg ctagttgaac
     6001 gcttccatct tcaatgttgt gtctaatttt gaagttaact ttgattccat tcttttgttt
     6061 gtctgccatg atgtatacat tgtgtgagtt atagttgtat tccaatttgt gtccaagaat
     6121 gtttccatct tctttaaaat caataccttt taactcgatt ctattaacaa gggtatcacc
     6181 ttcaaacttg acttcagcac gtgtcttgta gttcccgtca tctttgaaaa atatagttct
     6241 ttcctgtacg taaccttcgg gcatggcact cttgaaaaag tcatgctgtt tcatgtgatc
     6301 tgggtatctc gcaaagcatt gaacaccata accgaaagta gtgacaagtg ttggccacgg
     6361 aacaggtagt tttccagtag tgcaaataaa tttaagggta agttttccgt atgttgcatc
     6421 accttcaccc tctccactga cagaaaattt gtgcccatta acatcaccat ctaattcaac
     6481 aagaattggg acaactccag tgaaaagttc ttctccttta cgcatattat acctccttaa
     6541 tttagtgtca tctacaactg gcatgcgcgg ttatcctctg ttataagctt tctccagagc
     6601 cagataacgc cgctgtctct ggattgccga aatatgcttc ccgctacctg ggaaggggct
     6661 atcaactgta ctgcacggta atgtgacgtc ctttgcatac atgcagtaca tcaatgtatt
     6721 actgtagcat cctgactgtt ttagcatagc tttcgctttg tgtctcctgg tgtctcgctt
     6781 cagcatgacc caggtcgcct tccgttgcgc gatttggtta gtacgcgtac tctgtcagga
     6841 aaattgacgc agtggagtag caaaacctag ggaccgtctc gagagaatca atattaatcc
     6901 aacgcgtggc atcaaataaa acgaaaggct cagtcgaaag actgggcctt tcgttttatc
     6961 tgttgtttgt cggtgaacgc tctcctgagt aggacaaatc cgccgcccta gacttaggcg
     7021 ttcggctgcg gcgagcggta tcagctcact caaagaagtg gggcccagag agaatattca
     7081 ggccagttat gctttctggc ctgtaacaaa ggacattaag taaagacaga taaacgtaga
     7141 ctaaaacgtg gtcgcatcag ggtgctggct tttcaagttc cttaagaatg gcctcaattt
     7201 tctctataca ctcagttgga acacgggacc tgtccaggtt aagcaccatt ttatcgccct
     7261 tatacaatac tgtcgctcca ggagcaaact gatgtcgtga gcttaaacta gttcttgatg
     7321 cagatgacgt tttaagcaca gaagttaaaa gagtgataac ttcttcagct tcaaatatca
     7381 ccccagcttt tttctgctca tgaaggttag atgcctgctg cttaagtaat tcctctttat
     7441 ctgtaaaggc tttttgaagt gcatcacctg accgggcaga tagttcaccg gggtgagaaa
     7501 aaagagcaac aactgattta ggcaatttgg cggtgttgat acagcgggta ataatcttac
     7561 gtgaaatatt ttccgcatca gccagcgcag aaatatttcc agcaaattca ttctgcaatc
     7621 ggcttgcata acgctgacca cgttcataag cacttgttgg gcgataatcg ttacccaatc
     7681 tggataatgc agccatctgc tcatcatcca gctcgccaac cagaacacga taatcac
//
