LOCUS       sequence-141459-        6141 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pCM66T with shortened pHps2 promoter driving RFP expression (maxRFP 
            = BBa_E1010).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6141)
  TITLE     broad host range pcm66t
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6141)
  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            "Unpublished"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: pJM103: 150bp Hps2 RFP on pCM66T
FEATURES             Location/Qualifiers
     source          1..6141
                     /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             2167..2844
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     primer_bind     2942..2964
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2965..3780)
                     /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(2965..3780)
                     /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     3688..3707
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3772..3791
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             complement(4066..5214)
                     /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(5486..5857)
                     /codon_start=1
                     /gene="traJ"
                     /product="oriT-recognizing protein"
                     /label=traJ
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSL*AYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
                     KQDELGKVMMGVVRPRAEP"
     oriT            5890..5999
                     /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 ccaggccact ttattcaagt agtacataag
     2041 tgctattgga tttacctagc ttaacgttga catcaacttt gattccaact taagatagtt
     2101 tttctaaaaa ctaagcactt aattttttaa accaactgga ggaagtacat cgtggcatta
     2161 actcaaatgg caagtagtga agacgttatc aaagagttca tgcgtttcaa agttcgtatg
     2221 gaaggttccg ttaacggtca cgagttcgaa atcgaaggtg aaggtgaagg tcgtccgtac
     2281 gaaggtaccc agaccgctaa actgaaagtt accaaaggtg gtccgctgcc gttcgcttgg
     2341 gacatcctgt ccccgcagtt ccagtacggt tccaaagctt acgttaaaca cccggctgac
     2401 atcccggact acctgaaact gtccttcccg gaaggtttca aatgggaacg tgttatgaac
     2461 ttcgaagacg gtggtgttgt taccgttacc caggactcct ccctgcaaga cggtgagttc
     2521 atctacaaag ttaaactgcg tggtaccaac ttcccgtccg acggtccggt tatgcagaaa
     2581 aaaaccatgg gttgggaagc ttccaccgaa cgtatgtacc cggaagacgg tgctctgaaa
     2641 ggtgaaatca aaatgcgtct gaaactgaaa gacggtggtc actacgacgc tgaagttaaa
     2701 accacctaca tggctaaaaa accggttcag ctgccgggtg cttacaaaac cgacatcaaa
     2761 ctggacatca cctcccacaa cgaagactac accatcgttg aacagtacga acgtgctgaa
     2821 ggtcgtcact ccaccggtgc ttaataatta agccagcccc gacacccgcc aacacccgct
     2881 gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc
     2941 tccgggagct gcatgtgtca gaggttagaa aaactcatcg agcatcaaat gaaactgcaa
     3001 tttattcata tcaggattat caataccata tttttgaaaa agccgtttct gtaatgaagg
     3061 agaaaactca ccgaggcagt tccataggat ggcaagatcc tggtatcggt ctgcgattcc
     3121 gactcgtcca acatcaatac aacctattaa tttcccctcg tcaaaaataa ggttatcaag
     3181 tgagaaatca ccatgagtga cgactgaatc cggtgagaat ggcaaaagct tatgcatttc
     3241 tttccagact tgttcaacag gccagccatt acgctcgtca tcaaaatcac tcgcatcaac
     3301 caaaccgtta ttcattcgtg attgcgcctg agcgagacga aatacgcgat cgctgttaaa
     3361 aggacaatta caaacaggaa tcgaatgcaa ccggcgcagg aacactgcca gcgcatcaac
     3421 aatattttca cctgaatcag gatattcttc taatacctgg aatgctgttt tcccggggat
     3481 cgcagtggtg agtaaccatg catcatcagg agtacggata aaatgcttga tggtcggaag
     3541 aggcataaat tccgtcagcc agtttagtct gaccatctca tctgtaacat cattggcaac
     3601 gctacctttg ccatgtttca gaaacaactc tggcgcatcg ggcttcccat acaatcgata
     3661 gattgtcgca cctgattgcc cgacattatc gcgagcccat ttatacccat ataaatcagc
     3721 atccatgttg gaatttaatc gcggcctcga gcaagacgtt tcccgttgaa tatggctcat
     3781 aacacccctt gtattactgt ttatgtaagc agacagtttt attgttcatg atgatatatt
     3841 tttatcttgt gcaatgtaac atcagagatt ttgagacaca acgtggcttt cccccccccc
     3901 cctgcaggtc cgacacgggg atggatggcg ttcccgatca tggtcctgct tgcttcggga
     3961 gcgatactga gcgaagcaag tgcgtcgagc agtgcccgct tgttcctgaa atgccagtaa
     4021 agcgctggct gctgaacccc cagccggaac tgaccccaca aggccctagc gtttgcaatg
     4081 caccaggtca tcattgaccc aggcgtgttc caccaggccg ctgcctcgca actcttcgca
     4141 ggcttcgccg acctgctcgc gccacttctt cacgcgggtg gaatccgatc cgcacatgag
     4201 gcggaaggtt tccagcttga gcgggtacgg ctcccggtgc gagctgaaat agtcgaacat
     4261 ccgtcgggcc gtcggcgaca gcttgcggta cttctcccat atgaatttcg tgtagtggtc
     4321 gccagcaaac agcacgacga tttcctcgtc gatcaggacc tggcaacggg acgttttctt
     4381 gccacggtcc aggacgcgga agcggtgcag cagcgacacc gattccaggt gcccaacgcg
     4441 gtcggacgtg aagcccatcg ccgtcgcctg taggcgcgac aggcattcct cggccttcgt
     4501 gtaataccgg ccattgatcg accagcccag gtcctggcaa agctcgtaga acgtgaaggt
     4561 gatcggctcg ccgatagggg tgcgcttcgc gtactccaac acctgctgcc acaccagttc
     4621 gtcatcgtcg gcccgcagct cgacgccggt gtaggtgatc ttcacgtcct tgttgacgtg
     4681 gaaaatgacc ttgttttgca gcgcctcgcg cgggattttc ttgttgcgcg tggtgaacag
     4741 ggcagagcgg gccgtgtcgt ttggcatcgc tcgcatcgtg tccggccacg gcgcaatatc
     4801 gaacaaggaa agctgcattt ccttgatctg ctgcttcgtg tgtttcagca acgcggcctg
     4861 cttggcctcg ctgacctgtt ttgccaggtc ctcgccggcg gtttttcgct tcttggtcgt
     4921 catagttcct cgcgtgtcga tggtcatcga cttcgccaaa cctgccgcct cctgttcgag
     4981 acgacgcgaa cgctccacgg cggccgatgg cgcgggcagg gcagggggag ccagttgcac
     5041 gctgtcgcgc tcgatcttgg ccgtagcttg ctggaccatc gagccgacgg actggaaggt
     5101 ttcgcggggc gcacgcatga cggtgcggct tgcgatggtt tcggcatcct cggcggaaaa
     5161 ccccgcgtcg atcagttctt gcctgtatgc cttccggtca aacgtccgat tcattcaccc
     5221 tccttgcggg attgccccga ctcacgccgg ggcaatgtgc ccttattcct gatttgaccc
     5281 gcctggtgcc ttggtgtcca gataatccac cttatcggca atgaagtcgg tcccgtagac
     5341 cgtctggccg tccttctcgt acttggtatt ccgaatcttg ccctgcacga ataccagctc
     5401 cgcgaagtcg ctcttcttga tggagcgcat ggggacgtgc ttggcaatca cgcgcacccc
     5461 ccggccgttt tagcggctaa aaaagtcatg gctctgccct cgggcggacc acgcccatca
     5521 tgaccttgcc aagctcgtcc tgcttctctt cgatcttcgc cagcagggcg aggatcgtgg
     5581 catcaccgaa ccgcgccgtg cgcgggtcgt cggtgagcca gagtttcagc aggccgccca
     5641 ggcggcccag gtcgccattg atgcgggcca gctcgcggac gtgctcatag tccacgacgc
     5701 ccgtgatttt gtagccctgg ccgacggcca gcaggtaggc ctacaggctc atgccggccg
     5761 ccgccgcctt ttcctcaatc gctcttcgtt cgtctggaag gcagtacacc ttgataggtg
     5821 ggctgccctt cctggttggc ttggtttcat cagccatccg cttgccctca tctgttacgc
     5881 cggcggtagc cggccagcct cgcagagcag gattcccgtt gagcaccgcc aggtgcgaat
     5941 aagggacagt gaagaaggaa cacccgctcg cgggtgggcc tacttcacct atcctgcccg
     6001 gctgacgccg ttggatacac caaggaaagt ctacacgaac cctttggcaa aatcctgtat
     6061 atcgtgcgaa aaaggatgga tataccgaaa aaatcgctat aatgaccccg aagcagggtt
     6121 atgcagcgga aaagatccgt c
//
