LOCUS       sequence-332751-        5671 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mono-Responsive Reporter Gene Constructs for triple red.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5671)
  AUTHORS   Kolbe K, Bell AC, Prosser GA, Assmann M, Yang HJ, Forbes HE, 
            Gallucci S, Mayer-Barber KD, Boshoff HI, Barry Iii CE
  TITLE     Development and Optimization of Chromosomally-Integrated Fluorescent
            Mycobacterium tuberculosis Reporter Constructs.
  JOURNAL   Front Microbiol. 2020 Dec 9;11:591866. doi: 
            10.3389/fmicb.2020.591866. eCollection 2020.
  PUBMED    33362741
REFERENCE   2  (bases 1 to 5671)
  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: "Front 
            Microbiol."; date: "2020-12-9"; volume: "11:591866. doi"; pages: " 
            10.3389/fmicb.2020.591866. eCollection 2020"
COMMENT     Sequence Label: pGilesSHdI_psmyc_His6_triple red
FEATURES             Location/Qualifiers
     source          1..5671
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             136..144
                     /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)"
     protein_bind    complement(597..630)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(682..1392)
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby2, with improved brightness and photostability (Bajar 
                     et al., 2016)"
                     /label=mRuby3
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGRPYEGVQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPADIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVTQDTSLEDGELVYNVKVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTD
                     IALKVDGGGHLHCNFVTTYRSKKTVGNIKMPGVHAVDHRLERIEESDNETYVVQREVAV
                     AKYSNLGGGMDELYK"
     CDS             complement(1405..1422)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             1438..1446
                     /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)"
     CDS             complement(2175..2192)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             2208..2216
                     /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)"
     CDS             complement(2239..2934)
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     2783..2801
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     CDS             complement(2950..2967)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             2983..2991
                     /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)"
     rep_origin      3531..4119
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4020..4039
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4273..4290
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    complement(5377..5410)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
ORIGIN
        1 gggtgggcga gtttgtcctg cgtgtgctcg gtcgagtagg ctctgggagt acccgtgtgt
       61 acgaccagca cggcatacat catttcgacg ccgagagatt cgccgcccga aatgagcacg
      121 atccgcatgc ttaattaagg aggtatctcc atggcagagc tcgaattcga agctttagct
      181 aattaattgg ggaccctaga ggtccccttt tttattttat cgataaaaaa aaaagccccg
      241 caatcgcggg gctttttttt ttacgcgtcg ccgacctaca cgtattcgca gagttaagcg
      301 gtcacccccg ccggtttgac cccaattttg accccaattg catcggggtt cggggcgatt
      361 cgccgatgtc cgccattgtg cgcaaccggg cgtgacctgc ggattcggta cccgggcgga
      421 tccacgcgga tgccttttga atgactgggg gtcaaggggt cgcaggttca aatcctgtca
      481 gcccgacgca gatcaggggc ggttttccgg agacgggaag ccgccccttt ttctgtttga
      541 ccccaatttt gaccccaatt gctccggcgt ccgtccgcgt ccgtcgaggg cgcgccataa
      601 cttcgtatag catacattat acgaagttat gatatcaaaa aaaagcccgc cattaggcgg
      661 gctcaattga gcagatcttt acttgtacag ctcgtccatg ccaccaccaa ggttgctgta
      721 tttggcaact gccacttctc tttgcactac gtaggtttca ttgtcactct cctcgatcct
      781 ttccaggcgg tgatcaacgg catggacacc gggcatcttg atgttcccga cggtcttttt
      841 tgacctgtaa gttgtcacga agttgcagtg cagatggcca ccaccatcaa ctttcagtgc
      901 gatgtcagtg tatcctctca gaccaccatc tgctggatac atcatctctg tattaggctc
      961 ccaacccttg gtcttcttct gcatcacggg accattggag ggaaagttta cccctctgac
     1021 cttgacgttg tagacgagct cgccatcctc aaggctggtg tcctgcgtga cggtgacgac
     1081 tccaccatct tcgtatctcg taactctttc ccaagtaaaa ccctcaggaa aggactgttt
     1141 aaagaaatca gggatgtcgg ccgggtactt gataaaggta cggctgccat acatgaacga
     1201 cgtggcaaga atgtcaaagg caaatggcag gggtcctccc tcgatgactt tgatcctcat
     1261 ggtttgcact ccctcgtacg gtctgccttc tccttcacct gtgcatttga attggtggcc
     1321 gttgaccgaa ccttccatga ccaccttcat acgcatattt tccttgatca gctcttcgcc
     1381 cttagacacc atggcgctac cgccgtggtg atggtgatgg tgaccgctca tggagatacc
     1441 tccttaatta aaacgataag ctttcactta tacaattcat ccatcccccc ggtggaatga
     1501 cgaccctccg agcgttcgta ttgttccacc accgtgtaat cttcgttatg gctcgtaata
     1561 tccagcttac ggtcgacatt atacgccccg ggcatttgca cgggtttttt ggctttgtag
     1621 gtggttttga agtccgccaa gtagcgccca ccatctttca accgcagggc cattttaatg
     1681 tcacccttca acacgccatc ttctggatac agccgctcgg tcgaggcctc ccagcccatc
     1741 gttttctttt gcatgacggg accgtcgggg gggaaattcg tcccccgcaa cttgacctta
     1801 tagatgaggg tcccgtcttc cagcgaggta tcttgggtga ccgtcaccgc acccccgtcc
     1861 tcaaaattca tcacacgttc ccatttgaac ccctcgggga agctctgttt gtagtagtcc
     1921 gggatgtcgg cggggtgctt ggtaaacgca cgcgaaccgt acatgaattg tgggctgagg
     1981 atatcccacg agaacgggag cgggccgcct ttcgtgacct tcaatttcgc ggtttgcgtc
     2041 ccttcgtatg gacggccctc gccctccccc tcgatttcaa actcatgccc gttcatcgac
     2101 ccttccatat gcactttgaa acgcataaac tctttaatca ccgcctcgcc cttgctcacc
     2161 atggcgctac cgccgtggtg atggtgatgg tgaccgctca tggagatacc tccttaatta
     2221 gcatgcggaa agctttcact tgtacagctc gtccatgccg ccggtggagt ggcggccctc
     2281 ggagcgttcg tactgttcca ccacggtgta gtcctcgttg tgggaggtga tgtccaactt
     2341 gcggtcgacg ttgtaggcgc cgggcatctg cacgggcttc ttggccttgt aggtggtctt
     2401 gaagtccgcc aggtagcggc cgccgtcctt caggcgcagg gccatcttaa tgtcgccctt
     2461 cagcacgccg tcctcggggt acaaccgctc ggtggacgct tcccagccca ttgtcttctt
     2521 ctgcattacg gggccgtcag gagggaagtt ggtgccgcgg agcttcacct tgtagatcag
     2581 ggtgccgtcc tccagggagg tgtcctgggt cacggtcacg gcgccgccgt cctcgaagtt
     2641 catcacgcgc tcccacttga agccctcggg gaaggactgc ttatagtagt cggggatgtc
     2701 ggcggggtgc ttggtgaagg ccctggagcc gtacatgaac tgaggggaca ggatgtccca
     2761 ggagaagggc agggggccac ccttggtcac cttcagcttg gcggtctggg tgccctcgta
     2821 ggggcggccc tcgccctcgc cctcgatctc gaactcgtgg ccgttcatgg agccctccat
     2881 gtgcaccttg aaccgcatga actccttgat cactgcctcg cccttgctca ccatgggccc
     2941 gctaccgccg tggtgatggt gatggtgacc gctcatggag atacctcctt aattaagcat
     3001 gcggatcgtg ctcatttcgg gcggcgaatc tctcggcgtc gaaatgatgt atgccgtgct
     3061 ggtcgtacac acgggtactc ccagagccta ctcgaccgag cacacgcagg acaaactcgc
     3121 ccaccccggg tgggttctcc cgctcgtcag agacccttct cacaccgggc ctgtgcgtgc
     3181 gaacgcacag atcaacgccg ggatggcgta gagctcactg cgcagcaccg tggggatgcg
     3241 gccgatcatc acgtcgcgga tcgtgccgcc tcccacggcc gtgacggtgc cgacgatccg
     3301 ctagccgaga atgggagacc ctgcaccacg gcaatccaat aattgtagaa atattggtaa
     3361 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     3421 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     3481 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     3541 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     3601 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     3661 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
     3721 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     3781 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     3841 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     3901 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     3961 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     4021 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     4081 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     4141 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     4201 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
     4261 acgaccgagc gcagcgagtc agtgagcgag gaagccgacg gtatcgggta ccctcgagcc
     4321 ggccgtgcgg aattaagccg gcccgtaccc tgtgaataga ggtccgctgt gacacaagaa
     4381 tccctgttac ttctcgaccg tattgattcg gatgattcct acgcgagcct gcggaacgac
     4441 caggaattct gggagccgct ggcccgccga gccctggagg agctcgggct gccggtgccg
     4501 ccggtgctgc gggtgcccgg cgagagcacc aaccccgtac tggtcggcga gcccggcccg
     4561 gtgatcaagc tgttcggcga gcactggtgc ggtccggaga gcctcgcgtc ggagtcggag
     4621 gcgtacgcgg tcctggcgga cgccccggtg ccggtgcccc gcctcctcgg ccgcggcgag
     4681 ctgcggcccg gcaccggagc ctggccgtgg ccctacctgg tgatgagccg gatgaccggc
     4741 accacctggc ggtccgcgat ggacggcacg accgaccgga acgcgctgct cgccctggcc
     4801 cgcgaactcg gccgggtgct cggccggctg cacagggtgc cgctgaccgg gaacaccgtg
     4861 ctcacccccc attccgaggt cttcccggaa ctgctgcggg aacgccgcgc ggcgaccgtc
     4921 gaggaccacc gcgggtgggg ctacctctcg ccccggctgc tggaccgcct ggaggactgg
     4981 ctgccggacg tggacacgct gctggccggc cgcgaacccc ggttcgtcca cggcgacctg
     5041 cacgggacca acatcttcgt ggacctggcc gcgaccgagg tcaccgggat cgtcgacttc
     5101 accgacgtct atgcgggaga ctcccgctac agcctggtgc aactgcatct caacgccttc
     5161 cggggcgacc gcgagatcct ggccgcgctg ctcgacgggg cgcagtggaa gcggaccgag
     5221 gacttcgccc gcgaactgct cgccttcacc ttcctgcacg acttcgaggt gttcgaggag
     5281 accccgctgg atctctccgg cttcaccgat ccggaggaac tggcgcagtt cctctggggg
     5341 ccgccggaca ccgcccccgg cgcctgacgc cccgggataa cttcgtatag catacattat
     5401 acgaagttat atgcattgac taagctagat acctgaaaaa aaaaaagcgc cgcagttgcg
     5461 gcgctttttt ttttctcgac tagtgtttaa actctagagg atcgtcggca ccgtcacggc
     5521 cgtgggaggc ggcacgatcc gcgacgtgat gatcggccgc atccccacgg tgctgcgcag
     5581 tgagctctac gccatcccgg cgttgatctg tgcgttcgca cgcacaggcc cggtgtgaga
     5641 agggtctctg acgagcggga gaacccaccc g
//
