LOCUS       sequence-356596-        4493 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  aTc-inducible expression of dimerized mNeonGreen (mNG-mNG).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4493)
  AUTHORS   Butcher RJ, Tabor JJ
  TITLE     Real-time detection of response regulator phosphorylation dynamics 
            in live bacteria.
  JOURNAL   Proc Natl Acad Sci U S A. 2022 Aug 30;119(35):e2201204119. doi: 
            10.1073/pnas.2201204119. Epub 2022 Aug 22.
  PUBMED    35994658
REFERENCE   2  (bases 1 to 4493)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2022-08-30"; volume: "119(35):e2201204119. 
            doi"; pages: " 10.1073/pnas.2201204119. Epub 2022 Aug 22"
COMMENT     Sequence Label: pRB121
FEATURES             Location/Qualifiers
     source          1..4493
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(66..654)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(146..165)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(742..836)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(857..1516)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAKCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     1431..1450
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     primer_bind     1685..1703
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(1723..1741,1742..1747,1748..1764,1765..1770,
                     1771..1796)
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    1723..1741
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1748..1766
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     terminator      3339..3410
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3426..3453
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     promoter        3612..3646
                     /label=BBa_J23106
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.47"
     CDS             3681..4307
                     /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(3757..3776)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     terminator      4313..4399
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
       61 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
      121 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
      181 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
      241 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
      301 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
      361 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
      421 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
      481 tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca
      541 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
      601 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
      661 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
      721 ttggtcatga ctagtgcttg gattctcacc aataaaaaac gcccggcggc aaccgagcgt
      781 tctgaacaaa tccagatgga gttctgaggt cattactgga tctatcaaca ggagtccaag
      841 cgagctcgat atcaaattac gccccgccct gccactcatc gcagtactgt tgtaattcat
      901 taagcattct gccgacatgg aagccatcac aaacggcatg atgaacctga atcgccagcg
      961 gcatcagcac cttgtcgcct tgcgtataat atttgcccat ggtgaaaacg ggggcgaaga
     1021 agttgtccat attggccacg tttaaatcaa aactggtgaa actcacccag ggattggctg
     1081 agacgaaaaa catattctca ataaaccctt tagggaaata ggccaggttt tcaccataac
     1141 acgccacatc ttgcgaatat atgtgtagaa actgccggaa atcgtcgtgg tattcactcc
     1201 agagcgatga aaacgtttca gtttgctcat ggaaaacggt gtaacaaggg tgaacactat
     1261 cccatatcac cagctcaccg tctttcattg ccatacgaaa ttccggatga gcattcatca
     1321 ggcgggcaag aatgtgaata aaggccggat aaaacttgtg cttatttttc tttacggtct
     1381 ttaaaaaggc cgtaatatcc agctgaacgg tctggttata ggtacattta gcaactgact
     1441 gaaatgcctc aaaatgttct ttacgatgcc attgggatat atcaacggtg gtatatccag
     1501 tgattttttt ctccatttta gcttccttag ctcctgaaaa tctcgataac tcaaaaaata
     1561 cgaccggtag tgatcttatt tcattatggt gaaagttgga acctcttacg ggccggccaa
     1621 cgtctcattt tcgccagata tcgacgtcta agaaaccatt attatcatga cattaaccta
     1681 taaaaatagg cgtatcacga ggccctttcg tcttcacctc gatccctatc agtgatagag
     1741 attgacatcc ctatcagtga tagagatact gagcacatca gcaggacgca ctgaccgggc
     1801 ccaagttcac ttaaaaagga gatcaacaat gaaagcaatt ttcgtactga aacatcttaa
     1861 tcatgcctag gaagttttct aatggtttcg aaaggtgagg gggataacat ggcgtcgctg
     1921 cctgctacac atgaactgca tattttcggc agcatcaacg gagttgattt cgacatggtg
     1981 ggccaaggca ccggcaaccc gaacgacggc tatgaagaat tgaatttgaa atctaccaag
     2041 ggcgacttgc aatttagtcc gtggattctg gtaccgcata ttggttatgg ttttcatcag
     2101 tatctgccct acccagacgg aatgagccca tttcaggcgg ctatggtgga tggcagtggc
     2161 tatcaggtcc atcgcaccat gcaattcgag gatggggcga gcctgaccgt gaattatcgg
     2221 tacacttacg aaggttcaca tattaaaggt gaagcacaag ttaaaggcac cggcttcccg
     2281 gcggatggcc cagtgatgac caacagcctg acggcggcgg actggtgtcg tagcaaaaag
     2341 acgtatccca atgataaaac aattatttct acctttaaat ggagttacac cacgggtaac
     2401 ggtaaacgct accgtagcac ggctcgcacc acatacacct ttgcgaaacc catggccgcg
     2461 aactatctga aaaatcagcc gatgtatgtg tttcgcaaaa cggaactgaa acattcgaaa
     2521 actgaactca atttcaaaga gtggcaaaaa gccttcaccg acgtaatggg catggatgaa
     2581 ctgtacaaag gtggcggtgg ctctggaggt ggtgggtcca tggtttcgaa aggtgagggg
     2641 gataacatgg cgtcgctgcc tgctacacat gaactgcata ttttcggcag catcaacgga
     2701 gttgatttcg acatggtggg ccaaggcacc ggcaacccga acgacggcta tgaagaattg
     2761 aatttgaaat ctaccaaggg cgacttgcaa tttagtccgt ggattctggt accgcatatt
     2821 ggttatggtt ttcatcagta tctgccctac ccagacggaa tgagcccatt tcaggcggct
     2881 atggtggatg gcagtggcta tcaggtccat cgcaccatgc aattcgagga tggggcgagc
     2941 ctgaccgtga attatcggta cacttacgaa ggttcacata ttaaaggtga agcacaagtt
     3001 aaaggcaccg gcttcccggc ggatggccca gtgatgacca acagcctgac ggcggcggac
     3061 tggtgtcgta gcaaaaagac gtatcccaat gataaaacaa ttatttctac ctttaaatgg
     3121 agttacacca cgggtaacgg taaacgctac cgtagcacgg ctcgcaccac atacaccttt
     3181 gcgaaaccca tggccgcgaa ctatctgaaa aatcagccga tgtatgtgtt tcgcaaaacg
     3241 gaactgaaac attcgaaaac tgaactcaat ttcaaagagt ggcaaaaagc cttcaccgac
     3301 gtaatgggca tggatgaact gtacaaatga ccaggcatca aataaaacga aaggctcagt
     3361 cgaaagactg ggcctttcgt tttatctgtt gtttgtcggt gaacgctctc tactagagtc
     3421 acactggctc accttcgggt gggcctttct gcgtttatag taaatcactg cataattcgt
     3481 gtcgctcaag gcgcactccc gttctggata atgttttttg cgccgacatc ataacggttc
     3541 tggcaaatat tctgaaatga gctgctcgta cccgtggatc ctctagagtc gacctgcagg
     3601 catgcaagct ttttacggct agctcagtcc taggtatagt gctagcccag ccagagaaac
     3661 actctttaac agggggtagt atgatgtctc gtttagataa aagtaaagtg attaacagcg
     3721 cattagagct gcttaatgag gtcggaatcg aaggtttaac aacccgtaaa ctcgcccaga
     3781 agctaggtgt agagcagcct acattgtatt ggcatgtaaa aaataagcgg gctttgctcg
     3841 acgccttagc cattgagatg ttagataggc accatactca cttttgccct ttagaagggg
     3901 aaagctggca agatttttta cgtaataacg ctaaaagttt tagatgtgct ttactaagtc
     3961 atcgcgatgg agcaaaagta catttaggta cacggcctac agaaaaacag tatgaaactc
     4021 tcgaaaatca attagccttt ttatgccaac aaggtttttc actagagaat gcattatatg
     4081 cactcagcgc agtggggcat tttactttag gttgcgtatt ggaagatcaa gagcatcaag
     4141 tcgctaaaga agaaagggaa acacctacta ctgatagtat gccgccatta ttacgacaag
     4201 ctatcgaatt atttgatcac caaggtgcag agccagcctt cttattcggc cttgaattga
     4261 tcatatgcgg attagaaaaa caacttaaat gtgaaagtgg gtcttaaggc atcaaataaa
     4321 acgaaaggct cagtcgaaag actgggcctt tcgttttatc tgttgtttgt cggtgaacgc
     4381 tctcctgagt aggacaaatc cgccgcccta gacctagggc gttcggctgc ggcgagcggt
     4441 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acg
//
