LOCUS       sequence-308758-        5184 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  tdTomato, StrepR, L5int, attP to create RFP mycobacteria.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5184)
  AUTHORS   Meijers AS, Troost R, Ummels R, Maaskant J, Speer A, Nejentsev S, 
            Bitter W, Kuijl CP
  TITLE     Efficient genome editing in pathogenic mycobacteria using 
            Streptococcus thermophilus CRISPR1-Cas9.
  JOURNAL   Tuberculosis (Edinb). 2020 Sep;124:101983. doi: 
            10.1016/j.tube.2020.101983. Epub 2020 Aug 12.
  PUBMED    32829077
REFERENCE   2  (bases 1 to 5184)
  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: 
            "Tuberculosis (Edinb)."; date: "2020-09"; volume: "124:101983. doi";
            pages: " 10.1016/j.tube.2020.101983. Epub 2020 Aug 12"
COMMENT     Sequence Label: pTdTomato-L5
FEATURES             Location/Qualifiers
     source          1..5184
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             235..1026
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      1356..1944
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1845..1864
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             4359..5060
                     /codon_start=1
                     /product="dimeric variant of DsRed fluorescent protein 
                     (Shaner et al., 2004)"
                     /label=dTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLYGMDELYK"
     primer_bind     complement(4492..4510)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     4760..4779
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     4954..4977
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     terminator      5116..5162
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 gctggggggg aaagccacgt tgtgtctcaa aatctctgat gttacattgc acaagataaa
       61 aatatatcat catgaacaat aaaactgtct gcttacataa acagtaatag ggtacagtct
      121 atgcctcggg catccaagca gcaagcgcgt tacgccgtgg gtcgatgttt gatgttatgg
      181 agcagcaacg atgttacgca gcagggcagt cgccctaaaa caaagttaaa catcatgagg
      241 gaagcggtga tcgccgaagt atcgactcaa ctatcagagg tagttggcgt catcgagcgc
      301 catctcgaac cgacgttgct ggccgtacat ttgtacggct ccgcagtgga tggcggcctg
      361 aagccacaca gtgatattga tttgctggtt acggtgaccg taaggcttga tgaaacaacg
      421 cggcgagctt tgatcaacga ccttttggaa acttcggctt cccctggaga gagcgagatt
      481 ctccgcgctg tagaagtcac cattgttgtg cacgacgaca tcattccgtg gcgttatcca
      541 gctaagcgcg aactgcaatt tggagaatgg cagcgcaatg acattcttgc aggtatcttc
      601 gagccagcca cgatcgacat tgatctggct atcttgctga caaaagcaag agaacatagc
      661 gttgccttgg taggtccagc ggcggaggaa ctctttgatc cggttcctga acaggatcta
      721 tttgaggcgc taaatgaaac cttaacgcta tggaactcgc cgcccgactg ggctggcgat
      781 gagcgaaatg tagtgcttac gttgtcccgc atttggtaca gcgcagtaac cggcaaaatc
      841 gcgccgaagg atgtcgctgc cgactgggca atggagcgcc tgccggccca gtatcagccc
      901 gtcatacttg aagctagaca ggcttatctt ggacaagaag aagatcgctt ggcctcgcgc
      961 gcagatcagt tggaagaatt tgtccactac gtgaaaggcg agatcaccaa ggtagtcggc
     1021 aaataatcct agttccatga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
     1081 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
     1141 tggtttgttt gccggatcaa gcgtggcaaa gcaaaagttc aaaatcacca actctagaca
     1201 ggcttatctt ggacaagaag aagatcgctt ggcctcgcgc gcagatcagt tggaagaatt
     1261 tgtccactac gtgaaaggcg agatcaccaa ggtagtcggc aaataatcct agttccatga
     1321 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     1381 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     1441 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     1501 gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     1561 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     1621 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     1681 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     1741 cgtgagctat gagaaagcgc cacgcttccc gaggggagaa aggcggacag gtatccggta
     1801 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     1861 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     1921 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     1981 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     2041 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcaac
     2101 gcgtgcggcc gcggtacccg gggatcctct agagtcgacc accaagggca ccatctctgc
     2161 ttgggccacc ccgttggccg cagccagctc gctgagagcc gtgaacgaca gggcgaacgc
     2221 cagcccgccg acggcgaggg ttccgaccgc tgcaactccc ggtgcaacct tgtcccggtc
     2281 tattctcttc actgcaccag ctccaatctg gtgtgaatgc ccctcgtctg ttcgcgcagg
     2341 cggggggctc tattcgtttg tcagcatcga aagtagccag atcagggatg cgttgcaacc
     2401 gcgtatgccc aggtcagaag agtcgcacaa gagttgcaga cccctggaaa gaaaaatggc
     2461 cagagggcga aaacaccctc tgaccagcgg agcgggcgac gggaatcgaa cccgcgtagc
     2521 tagtttggaa gaatgggtgt ctgccgacca catatgggcc ggtcaagata ggtttttacc
     2581 ccctctcggc tgcatcctct aagtggaaag aaattgcagg tcgtagaagc gcgttgaagc
     2641 ctgagagttg cacaggagtt gcaacccggt agccttgttc acgacgagag gagacctagt
     2701 tggcacgtcg cggatgggga tcgctgaaga ctcagcgcag cgggaggatc caagcctcat
     2761 acgtcaaccc gcaggacggt gtgaggtact acgcgctgca gacctacgac aacaagatgg
     2821 acgccgaagc ctggctcgcg ggcgagaagc ggctcatcga gatggagacc tggacccctc
     2881 cacaggaccg ggcgaagaag gcagccgcca gcgccatcac gctggaggag tacacccgga
     2941 agtggctcgt ggagcgcgac ctcgcagacg gcaccaggga tctgtacagc gggcacgcgg
     3001 agcgccgcat ctacccggtg ctaggtgaag tggcggtcac agagatgacg ccagctctgg
     3061 tgcgtgcgtg gtgggccggg atgggtagga agcacccgac tgcccgccgg catgcctaca
     3121 acgtcctccg ggcggtgatg aacacagcgg tcgaggacaa gctgatcgca gagaacccgt
     3181 gccggatcga gcagaaggca gccgatgagc gcgacgtaga ggcgctgacg cctgaggagc
     3241 tggacatcgt cgccgctgag atcttcgagc actaccggat cgcggcatac atcctggcgt
     3301 ggacgagcct ccggttcgga gagctgatcg agcttcgccg caaggacatc gtggacgacg
     3361 gcatgacgat gaagctccgg gtgcgccgtg gcgcttcccg cgtggggaac aagatcgtcg
     3421 ttggcaacgc caagaccgtc cggtcgaagc gtcctgtgac ggttccgcct cacgtcgcgg
     3481 agatgatccg agcgcacatg aaggaccgta cgaagatgaa caagggcccc gaggcattcc
     3541 tggtgaccac gacgcagggc aaccggctgt cgaagtccgc gttcaccaag tcgctgaagc
     3601 gtggctacgc caagatcggt cggccggaac tccgcatcca cgacctccgc gctgtcggcg
     3661 ctacgttcgc cgctcaggca ggtgcgacga ccaaggagct gatggcccgt ctcggtcaca
     3721 cgactcctag gatggcgatg aagtaccaga tggcgtctga ggcccgcgac gaggctatcg
     3781 ctgaggcgat gtccaagctg gccaagacct cctgaaacgc aaaaagcccc cctcccaagg
     3841 acactgagtc ctaaagaggg gggtttcttg tcagtacgcg aagaaccacg cctggccgcg
     3901 agcgccagca ccgccgctct gtgcggagac ctgggcacca gccccgccgc cgccaggagc
     3961 attgccgttc ccgccagaaa tctagacggt gaccacaacg cgcccgcttt gatcggggac
     4021 gtctgcggcc gaccatttac gggtcttgtt gtcgttggcg gtcatgggcc gaacatactc
     4081 acccggatcg gagggccgag gacaaggtcg aacgaggggc atgacccggt gcggggcttc
     4141 ttgcactcgg cataggcgag tgctaagaat aacgttggca ctcgcgaccg gtgagtgcta
     4201 ggtcgggacg gtgaggccag gcccgtcgtc gcagcgagtg gcagcgagga caacttgagc
     4261 cgtccgtcgc gggcactgcg cccggccagc gtaagtagcg gggttgccgt cacccggtga
     4321 cccccgtttc atccccgatc cggaggaaag aattcataat ggtgagcaag ggcgaggagg
     4381 tcatcaaaga gttcatgcgc ttcaaggtgc gcatggaggg ctccatgaac ggccacgagt
     4441 tcgagatcga gggcgagggc gagggccgcc cctacgaggg cacccagacc gccaagctga
     4501 aggtgaccaa gggcggcccc ctgcccttcg cctgggacat cctgtccccc cagttcatgt
     4561 acggctccaa ggcgtacgtg aagcaccccg ccgacatccc cgattacaag aagctgtcct
     4621 tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga ggacggcggt ctggtgaccg
     4681 tgacccagga ctcctccctg caggacggca cgctgatcta caaggtgaag atgcgcggca
     4741 ccaacttccc ccccgacggc cccgtaatgc agaagaagac catgggctgg gaggcctcca
     4801 ccgagcgcct gtacccccgc gacggcgtgc tgaagggcga gatccaccag gccctgaagc
     4861 tgaaggacgg cggccactac ctggtggagt tcaagaccat ctacatggcc aagaagcccg
     4921 tgcaactgcc cggctactac tacgtggaca ccaagctgga catcacctcc cacaacgagg
     4981 actacaccat cgtggaacag tacgagcgct ccgagggccg ccaccacctg ttcctgtacg
     5041 gcatggacga gctgtacaag taaaagctta tcgatgtcga cgtagttaac tagcgtacga
     5101 tcgactgcca ggcatcaaat aaaacgaaag gctcagtcga aagactgggc ctttcgtttt
     5161 atgccatcat ggccgcggta atca
//
