LOCUS       sequence-141458-        6241 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pCM66T with promoter pHps2 driving RFP expression (maxRFP = 
            BBa_E1010).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6241)
  TITLE     broad host range pcm66t
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6241)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pJM102: pHps2 RFP on pCM66T
FEATURES             Location/Qualifiers
     source          1..6241
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      11..642
                     /label=oriV
                     /note="incP origin of replication"
     rep_origin      1105..1693
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1594..1613
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1847..1864
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1981..2002
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     CDS             2267..2944
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     primer_bind     3042..3064
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3065..3880)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(3065..3880)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     3788..3807
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3872..3891
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             complement(4166..5314)
                     /codon_start=1
                     /product="trans-acting replication protein that binds to 
                     and activates oriV"
                     /label=trfA
                     /translation="MNRTFDRKAYRQELIDAGFSAEDAETIASRTVMRAPRETFQSVGS
                     MVQQATAKIERDSVQLAPPALPAPSAAVERSRRLEQEAAGLAKSMTIDTRGTMTTKKRK
                     TAGEDLAKQVSEAKQAALLKHTKQQIKEMQLSLFDIAPWPDTMRAMPNDTARSALFTTR
                     NKKIPREALQNKVIFHVNKDVKITYTGVELRADDDELVWQQVLEYAKRTPIGEPITFTF
                     YELCQDLGWSINGRYYTKAEECLSRLQATAMGFTSDRVGHLESVSLLHRFRVLDRGKKT
                     SRCQVLIDEEIVVLFAGDHYTKFIWEKYRKLSPTARRMFDYFSSHREPYPLKLETFRLM
                     CGSDSTRVKKWREQVGEACEELRGSGLVEHAWVNDDLVHCKR"
     CDS             complement(5586..5957)
                     /codon_start=1
                     /gene="traJ"
                     /product="oriT-recognizing protein"
                     /label=traJ
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSL*AYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
                     KQDELGKVMMGVVRPRAEP"
     oriT            5990..6099
                     /note="incP origin of transfer"
ORIGIN
        1 gaccctttcc gacgctcacc gggctggttg ccctcgccgc tgggctggcg gccgtctatg
       61 gccctgcaaa cgcgccagaa acgccgtcga agccgtgtgc gagacaccgc ggccgccggc
      121 gttgtggata cctcgcggaa aacttggccc tcactgacag atgaggggcg gacgttgaca
      181 cttgaggggc cgactcaccc ggcgcggcgt tgacagatga ggggcaggct cgatttcggc
      241 cggcgacgtg gagctggcca gcctcgcaaa tcggcgaaaa cgcctgattt tacgcgagtt
      301 tcccacagat gatgtggaca agcctgggga taagtgccct gcggtattga cacttgaggg
      361 gcgcgactac tgacagatga ggggcgcgat ccttgacact tgaggggcag agtgctgaca
      421 gatgaggggc gcacctattg acatttgagg ggctgtccac aggcagaaaa tccagcattt
      481 gcaagggttt ccgcccgttt ttcggccacc gctaacctgt cttttaacct gcttttaaac
      541 caatatttat aaaccttgtt tttaaccagg gctgcgccct gtgcgcgtga ccgcgcacgc
      601 cgaagggggg tgccccccct tctcgaaccc tcccggcccg ctaacgcggg cctcccatcc
      661 ccccaggggc tgcgcccctc ggccgcgaac ggcctcaccc caaaaatggc agccaagctg
      721 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
      781 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
      841 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
      901 tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat
      961 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     1021 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     1081 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     1141 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     1201 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     1261 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     1321 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     1381 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     1441 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     1501 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     1561 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     1621 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     1681 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     1741 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     1801 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     1861 cgaggaagcg gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca
     1921 ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat
     1981 taatgtgagt tagctcactc attaggcacc ccaggcggtt tcgtggtaga ccaaaacaag
     2041 ctggaaaagg cactggctga aaagctgtaa tattgctgac ttgaatagcg ctaataatca
     2101 atttattagc gctattcact ttattcaagt agtacataag tgctattgga tttacctagc
     2161 ttaacgttga catcaacttt gattccaact taagatagtt tttctaaaaa ctaagcactt
     2221 aattttttaa accaactgga ggaagtacat cgtggcatta actcaaatgg caagtagtga
     2281 agacgttatc aaagagttca tgcgtttcaa agttcgtatg gaaggttccg ttaacggtca
     2341 cgagttcgaa atcgaaggtg aaggtgaagg tcgtccgtac gaaggtaccc agaccgctaa
     2401 actgaaagtt accaaaggtg gtccgctgcc gttcgcttgg gacatcctgt ccccgcagtt
     2461 ccagtacggt tccaaagctt acgttaaaca cccggctgac atcccggact acctgaaact
     2521 gtccttcccg gaaggtttca aatgggaacg tgttatgaac ttcgaagacg gtggtgttgt
     2581 taccgttacc caggactcct ccctgcaaga cggtgagttc atctacaaag ttaaactgcg
     2641 tggtaccaac ttcccgtccg acggtccggt tatgcagaaa aaaaccatgg gttgggaagc
     2701 ttccaccgaa cgtatgtacc cggaagacgg tgctctgaaa ggtgaaatca aaatgcgtct
     2761 gaaactgaaa gacggtggtc actacgacgc tgaagttaaa accacctaca tggctaaaaa
     2821 accggttcag ctgccgggtg cttacaaaac cgacatcaaa ctggacatca cctcccacaa
     2881 cgaagactac accatcgttg aacagtacga acgtgctgaa ggtcgtcact ccaccggtgc
     2941 ttaataatta agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg
     3001 tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca
     3061 gaggttagaa aaactcatcg agcatcaaat gaaactgcaa tttattcata tcaggattat
     3121 caataccata tttttgaaaa agccgtttct gtaatgaagg agaaaactca ccgaggcagt
     3181 tccataggat ggcaagatcc tggtatcggt ctgcgattcc gactcgtcca acatcaatac
     3241 aacctattaa tttcccctcg tcaaaaataa ggttatcaag tgagaaatca ccatgagtga
     3301 cgactgaatc cggtgagaat ggcaaaagct tatgcatttc tttccagact tgttcaacag
     3361 gccagccatt acgctcgtca tcaaaatcac tcgcatcaac caaaccgtta ttcattcgtg
     3421 attgcgcctg agcgagacga aatacgcgat cgctgttaaa aggacaatta caaacaggaa
     3481 tcgaatgcaa ccggcgcagg aacactgcca gcgcatcaac aatattttca cctgaatcag
     3541 gatattcttc taatacctgg aatgctgttt tcccggggat cgcagtggtg agtaaccatg
     3601 catcatcagg agtacggata aaatgcttga tggtcggaag aggcataaat tccgtcagcc
     3661 agtttagtct gaccatctca tctgtaacat cattggcaac gctacctttg ccatgtttca
     3721 gaaacaactc tggcgcatcg ggcttcccat acaatcgata gattgtcgca cctgattgcc
     3781 cgacattatc gcgagcccat ttatacccat ataaatcagc atccatgttg gaatttaatc
     3841 gcggcctcga gcaagacgtt tcccgttgaa tatggctcat aacacccctt gtattactgt
     3901 ttatgtaagc agacagtttt attgttcatg atgatatatt tttatcttgt gcaatgtaac
     3961 atcagagatt ttgagacaca acgtggcttt cccccccccc cctgcaggtc cgacacgggg
     4021 atggatggcg ttcccgatca tggtcctgct tgcttcggga gcgatactga gcgaagcaag
     4081 tgcgtcgagc agtgcccgct tgttcctgaa atgccagtaa agcgctggct gctgaacccc
     4141 cagccggaac tgaccccaca aggccctagc gtttgcaatg caccaggtca tcattgaccc
     4201 aggcgtgttc caccaggccg ctgcctcgca actcttcgca ggcttcgccg acctgctcgc
     4261 gccacttctt cacgcgggtg gaatccgatc cgcacatgag gcggaaggtt tccagcttga
     4321 gcgggtacgg ctcccggtgc gagctgaaat agtcgaacat ccgtcgggcc gtcggcgaca
     4381 gcttgcggta cttctcccat atgaatttcg tgtagtggtc gccagcaaac agcacgacga
     4441 tttcctcgtc gatcaggacc tggcaacggg acgttttctt gccacggtcc aggacgcgga
     4501 agcggtgcag cagcgacacc gattccaggt gcccaacgcg gtcggacgtg aagcccatcg
     4561 ccgtcgcctg taggcgcgac aggcattcct cggccttcgt gtaataccgg ccattgatcg
     4621 accagcccag gtcctggcaa agctcgtaga acgtgaaggt gatcggctcg ccgatagggg
     4681 tgcgcttcgc gtactccaac acctgctgcc acaccagttc gtcatcgtcg gcccgcagct
     4741 cgacgccggt gtaggtgatc ttcacgtcct tgttgacgtg gaaaatgacc ttgttttgca
     4801 gcgcctcgcg cgggattttc ttgttgcgcg tggtgaacag ggcagagcgg gccgtgtcgt
     4861 ttggcatcgc tcgcatcgtg tccggccacg gcgcaatatc gaacaaggaa agctgcattt
     4921 ccttgatctg ctgcttcgtg tgtttcagca acgcggcctg cttggcctcg ctgacctgtt
     4981 ttgccaggtc ctcgccggcg gtttttcgct tcttggtcgt catagttcct cgcgtgtcga
     5041 tggtcatcga cttcgccaaa cctgccgcct cctgttcgag acgacgcgaa cgctccacgg
     5101 cggccgatgg cgcgggcagg gcagggggag ccagttgcac gctgtcgcgc tcgatcttgg
     5161 ccgtagcttg ctggaccatc gagccgacgg actggaaggt ttcgcggggc gcacgcatga
     5221 cggtgcggct tgcgatggtt tcggcatcct cggcggaaaa ccccgcgtcg atcagttctt
     5281 gcctgtatgc cttccggtca aacgtccgat tcattcaccc tccttgcggg attgccccga
     5341 ctcacgccgg ggcaatgtgc ccttattcct gatttgaccc gcctggtgcc ttggtgtcca
     5401 gataatccac cttatcggca atgaagtcgg tcccgtagac cgtctggccg tccttctcgt
     5461 acttggtatt ccgaatcttg ccctgcacga ataccagctc cgcgaagtcg ctcttcttga
     5521 tggagcgcat ggggacgtgc ttggcaatca cgcgcacccc ccggccgttt tagcggctaa
     5581 aaaagtcatg gctctgccct cgggcggacc acgcccatca tgaccttgcc aagctcgtcc
     5641 tgcttctctt cgatcttcgc cagcagggcg aggatcgtgg catcaccgaa ccgcgccgtg
     5701 cgcgggtcgt cggtgagcca gagtttcagc aggccgccca ggcggcccag gtcgccattg
     5761 atgcgggcca gctcgcggac gtgctcatag tccacgacgc ccgtgatttt gtagccctgg
     5821 ccgacggcca gcaggtaggc ctacaggctc atgccggccg ccgccgcctt ttcctcaatc
     5881 gctcttcgtt cgtctggaag gcagtacacc ttgataggtg ggctgccctt cctggttggc
     5941 ttggtttcat cagccatccg cttgccctca tctgttacgc cggcggtagc cggccagcct
     6001 cgcagagcag gattcccgtt gagcaccgcc aggtgcgaat aagggacagt gaagaaggaa
     6061 cacccgctcg cgggtgggcc tacttcacct atcctgcccg gctgacgccg ttggatacac
     6121 caaggaaagt ctacacgaac cctttggcaa aatcctgtat atcgtgcgaa aaaggatgga
     6181 tataccgaaa aaatcgctat aatgaccccg aagcagggtt atgcagcgga aaagatccgt
     6241 c
//
