LOCUS       sequence-146948-        5990 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pCM66T RFP control vector: no promoter driving maxRFP (BBa_E1010).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5990)
  TITLE     broad host range pcm66t
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5990)
  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            "Unpublished"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: pJM100: no promoter RFP on pCM66T
FEATURES             Location/Qualifiers
     source          1..5990
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      11..642
                     /label=oriV
                     /note="incP origin of replication"
     rep_origin      1105..1692
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1593..1612
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1846..1863
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1980..2001
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     CDS             2016..2693
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     primer_bind     2791..2813
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2814..3629)
                     /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(2814..3629)
                     /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     3537..3556
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3621..3640
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             complement(3915..5063)
                     /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(5335..5706)
                     /codon_start=1
                     /gene="traJ"
                     /product="oriT-recognizing protein"
                     /label=traJ
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSL*AYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
                     KQDELGKVMMGVVRPRAEP"
     oriT            5739..5848
                     /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 tccttttttc tgcgcgtaat ctgctgcttg
     1141 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     1201 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
     1261 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     1321 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     1381 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     1441 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     1501 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     1561 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     1621 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     1681 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     1741 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
     1801 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc
     1861 gaggaagcgg aagagcgccc aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat
     1921 taatgcagct ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt
     1981 aatgtgagtt agctcactca ttaggcaccc caggcatggc aagtagtgaa gacgttatca
     2041 aagagttcat gcgtttcaaa gttcgtatgg aaggttccgt taacggtcac gagttcgaaa
     2101 tcgaaggtga aggtgaaggt cgtccgtacg aaggtaccca gaccgctaaa ctgaaagtta
     2161 ccaaaggtgg tccgctgccg ttcgcttggg acatcctgtc cccgcagttc cagtacggtt
     2221 ccaaagctta cgttaaacac ccggctgaca tcccggacta cctgaaactg tccttcccgg
     2281 aaggtttcaa atgggaacgt gttatgaact tcgaagacgg tggtgttgtt accgttaccc
     2341 aggactcctc cctgcaagac ggtgagttca tctacaaagt taaactgcgt ggtaccaact
     2401 tcccgtccga cggtccggtt atgcagaaaa aaaccatggg ttgggaagct tccaccgaac
     2461 gtatgtaccc ggaagacggt gctctgaaag gtgaaatcaa aatgcgtctg aaactgaaag
     2521 acggtggtca ctacgacgct gaagttaaaa ccacctacat ggctaaaaaa ccggttcagc
     2581 tgccgggtgc ttacaaaacc gacatcaaac tggacatcac ctcccacaac gaagactaca
     2641 ccatcgttga acagtacgaa cgtgctgaag gtcgtcactc caccggtgct taataattaa
     2701 gccagccccg acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg
     2761 catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttagaaa
     2821 aactcatcga gcatcaaatg aaactgcaat ttattcatat caggattatc aataccatat
     2881 ttttgaaaaa gccgtttctg taatgaagga gaaaactcac cgaggcagtt ccataggatg
     2941 gcaagatcct ggtatcggtc tgcgattccg actcgtccaa catcaataca acctattaat
     3001 ttcccctcgt caaaaataag gttatcaagt gagaaatcac catgagtgac gactgaatcc
     3061 ggtgagaatg gcaaaagctt atgcatttct ttccagactt gttcaacagg ccagccatta
     3121 cgctcgtcat caaaatcact cgcatcaacc aaaccgttat tcattcgtga ttgcgcctga
     3181 gcgagacgaa atacgcgatc gctgttaaaa ggacaattac aaacaggaat cgaatgcaac
     3241 cggcgcagga acactgccag cgcatcaaca atattttcac ctgaatcagg atattcttct
     3301 aatacctgga atgctgtttt cccggggatc gcagtggtga gtaaccatgc atcatcagga
     3361 gtacggataa aatgcttgat ggtcggaaga ggcataaatt ccgtcagcca gtttagtctg
     3421 accatctcat ctgtaacatc attggcaacg ctacctttgc catgtttcag aaacaactct
     3481 ggcgcatcgg gcttcccata caatcgatag attgtcgcac ctgattgccc gacattatcg
     3541 cgagcccatt tatacccata taaatcagca tccatgttgg aatttaatcg cggcctcgag
     3601 caagacgttt cccgttgaat atggctcata acaccccttg tattactgtt tatgtaagca
     3661 gacagtttta ttgttcatga tgatatattt ttatcttgtg caatgtaaca tcagagattt
     3721 tgagacacaa cgtggctttc cccccccccc ctgcaggtcc gacacgggga tggatggcgt
     3781 tcccgatcat ggtcctgctt gcttcgggag cgatactgag cgaagcaagt gcgtcgagca
     3841 gtgcccgctt gttcctgaaa tgccagtaaa gcgctggctg ctgaaccccc agccggaact
     3901 gaccccacaa ggccctagcg tttgcaatgc accaggtcat cattgaccca ggcgtgttcc
     3961 accaggccgc tgcctcgcaa ctcttcgcag gcttcgccga cctgctcgcg ccacttcttc
     4021 acgcgggtgg aatccgatcc gcacatgagg cggaaggttt ccagcttgag cgggtacggc
     4081 tcccggtgcg agctgaaata gtcgaacatc cgtcgggccg tcggcgacag cttgcggtac
     4141 ttctcccata tgaatttcgt gtagtggtcg ccagcaaaca gcacgacgat ttcctcgtcg
     4201 atcaggacct ggcaacggga cgttttcttg ccacggtcca ggacgcggaa gcggtgcagc
     4261 agcgacaccg attccaggtg cccaacgcgg tcggacgtga agcccatcgc cgtcgcctgt
     4321 aggcgcgaca ggcattcctc ggccttcgtg taataccggc cattgatcga ccagcccagg
     4381 tcctggcaaa gctcgtagaa cgtgaaggtg atcggctcgc cgataggggt gcgcttcgcg
     4441 tactccaaca cctgctgcca caccagttcg tcatcgtcgg cccgcagctc gacgccggtg
     4501 taggtgatct tcacgtcctt gttgacgtgg aaaatgacct tgttttgcag cgcctcgcgc
     4561 gggattttct tgttgcgcgt ggtgaacagg gcagagcggg ccgtgtcgtt tggcatcgct
     4621 cgcatcgtgt ccggccacgg cgcaatatcg aacaaggaaa gctgcatttc cttgatctgc
     4681 tgcttcgtgt gtttcagcaa cgcggcctgc ttggcctcgc tgacctgttt tgccaggtcc
     4741 tcgccggcgg tttttcgctt cttggtcgtc atagttcctc gcgtgtcgat ggtcatcgac
     4801 ttcgccaaac ctgccgcctc ctgttcgaga cgacgcgaac gctccacggc ggccgatggc
     4861 gcgggcaggg cagggggagc cagttgcacg ctgtcgcgct cgatcttggc cgtagcttgc
     4921 tggaccatcg agccgacgga ctggaaggtt tcgcggggcg cacgcatgac ggtgcggctt
     4981 gcgatggttt cggcatcctc ggcggaaaac cccgcgtcga tcagttcttg cctgtatgcc
     5041 ttccggtcaa acgtccgatt cattcaccct ccttgcggga ttgccccgac tcacgccggg
     5101 gcaatgtgcc cttattcctg atttgacccg cctggtgcct tggtgtccag ataatccacc
     5161 ttatcggcaa tgaagtcggt cccgtagacc gtctggccgt ccttctcgta cttggtattc
     5221 cgaatcttgc cctgcacgaa taccagctcc gcgaagtcgc tcttcttgat ggagcgcatg
     5281 gggacgtgct tggcaatcac gcgcaccccc cggccgtttt agcggctaaa aaagtcatgg
     5341 ctctgccctc gggcggacca cgcccatcat gaccttgcca agctcgtcct gcttctcttc
     5401 gatcttcgcc agcagggcga ggatcgtggc atcaccgaac cgcgccgtgc gcgggtcgtc
     5461 ggtgagccag agtttcagca ggccgcccag gcggcccagg tcgccattga tgcgggccag
     5521 ctcgcggacg tgctcatagt ccacgacgcc cgtgattttg tagccctggc cgacggccag
     5581 caggtaggcc tacaggctca tgccggccgc cgccgccttt tcctcaatcg ctcttcgttc
     5641 gtctggaagg cagtacacct tgataggtgg gctgcccttc ctggttggct tggtttcatc
     5701 agccatccgc ttgccctcat ctgttacgcc ggcggtagcc ggccagcctc gcagagcagg
     5761 attcccgttg agcaccgcca ggtgcgaata agggacagtg aagaaggaac acccgctcgc
     5821 gggtgggcct acttcaccta tcctgcccgg ctgacgccgt tggatacacc aaggaaagtc
     5881 tacacgaacc ctttggcaaa atcctgtata tcgtgcgaaa aaggatggat ataccgaaaa
     5941 aatcgctata atgaccccga agcagggtta tgcagcggaa aagatccgtc
//
