LOCUS       sequence-397042-        8468 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Broad host range plasmid for constitutive expression of mCherry. 
            Designed for expression in Pseudomonas aeruginosa..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8468)
  TITLE     Franklin lab plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8468)
  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            "Unpublished"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: pMF440
FEATURES             Location/Qualifiers
     source          1..8468
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(1186..1191,1192..1208,1209..1215)
                     /label=trc promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1223..1239
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     1228..1250
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1322..2032
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1467..1485)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1735..1754
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(2071..2088)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      2241..2327
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2419..2446
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        2465..2556
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2557..2625,2626..3417)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2775..2794)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3588..4176
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4077..4096
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4330..4347
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4362..4502
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4428..4447)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4588..4610
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4604..4795)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     6253..6272
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     oriT            6702..6811
                     /note="incP origin of transfer"
     CDS             complement(7581..8414)
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pRO1600 from Pseudomonas aeruginosa "
                     /label=pRO1600 Rep
                     /translation="MASPPMVYKSNALVEAAYRLSVQEQRIVLACISQVKRSEPVTDEV
                     MYSVTAEDIATMAGVPIESSYNQLKEAALRLKRREVRLTQEPNGKGKRPSVMITGWVQT
                     IIYREGEGRVELRFTKDMLPYLTELTKQFTKYALADVAKMDSTHAIRLYELLMQWDSIG
                     QREIEIDQLRKWFQLEGRYPSIKDFKLRVLDPAVTQINEHSPLQVEWAQRKTGRKVTHL
                     LFSFGPKKPAKAVGKAPAKRKAGKISDAEIAKQARPGETWEAARARLTQMPLDLA"
     rep_origin      join(8428..8468,1..311)
                     /label=pRO1600 oriV
                     /note="broad-host-range origin of replication from 
                     Pseudomonas aeruginosa plasmid pRO1600; requires the 
                     pRO1600 Rep protein for replication (West et al., 1994)"
ORIGIN
        1 aaatccgtcc ctttacaaca acaaatccgt cccttcttaa caacaaatcc gtcccttaat
       61 ggcaacaaat ccgtcccttt ttaaactcta caggccacgg attacgtggc ctgtagacgt
      121 cctaaaaggt ttaaaaggga aaaggaagaa aagggtggaa acgcaaaaaa cgcaccacta
      181 cgtggccccg ttggggccgc atttgtgccc ctgaaggggc gggggaggcg tctgggcaat
      241 ccccgtttta ccagtcccct atcgccgcct gagagggcgc aggaagcgag taatcagggt
      301 atcgaggcgg attcaccctt ggcgtccaac cagcggcacc agcggcgcct gagaggccta
      361 cagagcggtt ggacaccaag gggaggggct aagaccggtt tatcagtccc ctttccctcg
      421 tttctttcca acgcgatagc ccagcaaggc cgccaccgtt gccaccgtca ccccagcaag
      481 cacagccagt ggcgtgtaat tggggtctga cgctcagtgg aacgaaaact cacgttaagg
      541 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
      601 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
      661 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
      721 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
      781 gataccgcga gatccacgct caccggctcc agatttatca gcaataaacc agccagccgg
      841 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
      901 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
      961 tgctgcaggt cgacggatcc ggggaattct catgtttgac agcttatcat cgactgcacg
     1021 gtgcaccaat gcttctggcg tcaggcagcc atcggaagct gtggtatggc tgtgcaggtc
     1081 gtaaatcact gcataattcg tgtcgctcaa ggcgcactcc cgttctggat aatgtttttt
     1141 gcgccgacat cataacggtt ctggcaaata ttctgaaatg agctgttgac aattaatcat
     1201 ccggctcgta taatgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagacca
     1261 tggctgcagt ctagactcga gggtaccaga tctctgcagc caagcttagg agatcgatca
     1321 tatggtgagc aagggcgagg aggataacat ggccatcatc aaggagttca tgcgcttcaa
     1381 ggtgcacatg gagggctccg tgaacggcca cgagttcgag atcgagggcg agggcgaggg
     1441 ccgcccctac gagggcaccc agaccgccaa gctgaaggtg accaagggtg gccccctgcc
     1501 cttcgcctgg gacatcctgt cccctcagtt catgtacggc tccaaggcct acgtgaagca
     1561 ccccgccgac atccccgact acttgaagct gtccttcccc gagggcttca agtgggagcg
     1621 cgtgatgaac ttcgaggacg gcggcgtggt gaccgtgacc caggactcct ccctgcagga
     1681 cggcgagttc atctacaagg tgaagctgcg cggcaccaac ttcccctccg acggccccgt
     1741 aatgcagaag aagaccatgg gctgggaggc ctcctccgag cggatgtacc ccgaggacgg
     1801 cgccctgaag ggcgagatca agcagaggct gaagctgaag gacggcggcc actacgacgc
     1861 tgaggtcaag accacctaca aggccaagaa gcccgtgcag ctgcccggcg cctacaacgt
     1921 caacatcaag ttagacatca cctcccacaa cgaggactac accatcgtgg aacagtacga
     1981 acgcgccgag ggccgccact ccaccggcgg catggacgag ctgtacaagt agaagcttgg
     2041 ctgttttggc ggatgagaga agattttcag cctgatacag attaaatcag aacgcagaag
     2101 cggtctgata aaacagaatt tgcctggcgg cagtagcgcg gtggtcccac ctgaccccat
     2161 gccgaactca gaagtgaaac gccgtagcgc cgatggtagt gtggggtctc cccatgcgag
     2221 agtagggaac tgccaggcat caaataaaac gaaaggctca gtcgaaagac tgggcctttc
     2281 gttttatctg ttgtttgtcg gtgaacgctc tcctgagtag gacaaatccg ccgggagcgg
     2341 atttgaacgt tgcgaagcaa cggcccggag ggtggcgggc aggacgcccg ccataaactg
     2401 ccaggcatca aattaagcag aaggccatcc tgacggatgg cctttttgcg tttctacaaa
     2461 ctcttttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     2521 tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc
     2581 gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg
     2641 gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat
     2701 ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc
     2761 acttttaaag ttctgctatg tggcgcggta ttatcccgtg ttgacgccgg gcaagagcaa
     2821 ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa
     2881 aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt
     2941 gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct
     3001 tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat
     3061 gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg
     3121 cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg
     3181 atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt
     3241 attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg
     3301 ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg
     3361 gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg
     3421 tcagaccaag tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa
     3481 aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt
     3541 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt
     3601 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
     3661 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag
     3721 ataccaaata ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta
     3781 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
     3841 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
     3901 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
     3961 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
     4021 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
     4081 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
     4141 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
     4201 cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat
     4261 tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg
     4321 accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcctgatgcg gtattttctc
     4381 cttacgcatc tgtgcggtat ttcacaccgc atatggtgca ctctcagtac aatctgctct
     4441 gatgccgcat agttaagcca gtatacactc cgctatcgct acgtgactgg gtcatggctg
     4501 cgccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat
     4561 ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt
     4621 catcaccgaa acgcgcgagg cagctgcggt aaagctcatc agcgtggtcg tgaagcgatt
     4681 cacagatgtc tgcctgttca tccgcgtcca gctcgttgag tttctccaga agcgttaatg
     4741 tctggcttct gataaagcgg gccatgttaa gggcggtttt ttcctgtttg gtcactgatg
     4801 cctccgtgta agggggattt ctgttcatgg gggtaatgat accgatgaaa cgagagagga
     4861 tgctcacgat acgggttact gatgatgaac atgcccggtt actggaacgt tgtgagggta
     4921 aacaactggc ggtatggatg cggcgggacc agagaaaaat cactcagggt caatgccagc
     4981 gcttcgttaa tacagatgta ggtgttccac agggtagcca gcagcatcct gcgatgcaga
     5041 tccggaacat aatggtgcag ggcgctgact tccgcgtttc cagactttac gaaacacgga
     5101 aaccgaagac cattcatgtt gttgctcagg tcgcagacgt tttgcagcag cagtcgcttc
     5161 acgttcgctc gcgtatcggt gattcattct gctaaccagt aaggcaaccc cgccagccta
     5221 gccgggtcct caacgacagg agcacgatca tgcgcacccg tggccaggac ccaacgctgc
     5281 ccgagatgcg ccgcgtgcgg ctgctggaga tggcggacgc gatggatatg ttctgccaag
     5341 ggttggtttg cgcattcaca gttctccgca agaattgatt ggctccaatt cttggagtgg
     5401 tgaatccgtt agcgaggtgc cgccggcttc cattcaggtc gaggtggccc ggctccatgc
     5461 accgcgacgc aacgcgggga ggcagacaag gtatagggcg gcgcctacaa tccatgccaa
     5521 cccgttccat gtgctcgccg aggcggcata aatcgccgtg acgatcagcg gtccaatgat
     5581 cgaagttagg ctggtaagag ccgcgagcga tccttgaagc tgtccctgat ggtcgtcatc
     5641 tacctgcctg gacagcatgg cctgcaacgc gggcatcccg atgccgccgg aagcgagaag
     5701 aatcataatg gggaaggcca tccagcctcg cgtcgcgaac gccagcaaga cgtagcccag
     5761 cgcgtcggcc gccatgccgg cgataatggc ctgcttctcg ccgaaacgtt tggtggcggg
     5821 accagtgacg aaggcttgag cgagggcgtg caagattccg aataccgcaa gcgacaggcc
     5881 gatcatcgtc gcgctccagc gaaagcggtc ctcgccgaaa atgacccaga gcgctgccgg
     5941 cacctgtcct acgagttgca tgataaagaa gacagtcata agtgcggcga cgatagtcat
     6001 gccccgcgcc caccggaagg agctgactgg gttgaaggct ctcaagggca tcggtcgacg
     6061 ctctccctta tgcgactcct gcattaggaa gcagcccagt agtaggttga ggccgttgag
     6121 caccgccgcc gcaaggaatg gtgcatgcaa ggagatggcg cccaacagtc ccccggccac
     6181 ggggcctgcc accataccca cgccgaaaca agcgctcatg agcccgaagt ggcgagcccg
     6241 atcttcccca tcggtgatgt cggcgatata ggcgccagca accgcacctg tggcgccggt
     6301 gatgccggcc acgatgcgtc cggcgtagag gatcttccat acctaccagt tctgcgcctg
     6361 cagcaatggc aacaacgttg cccggatcgt ggcatcaccg aaccgcgccg tgcgcgggtc
     6421 gtcggtgagc cagagtttca gcaggccgcc caggcggccc aggtcgccat tgatgcgggc
     6481 cagctcgcgg acgtgctcat agtccacgac gcccgtgatt ttgtagccct ggccgacggc
     6541 cagcaggtag gccgacaggc tcatgccggc cgccgccgcc ttttcctcaa tcgctcttcg
     6601 ttcgtctgga aggcagtaca ccttgatagg tgggctgccc ttcctggttg gcttggtttc
     6661 atcagccatc cgcttgccct catctgttac gccggcggta gccggccagc ctcgcagagc
     6721 aggattcccg ttgagcaccg ccaggtgcga ataagggaca gtgaagaagg aacacccgct
     6781 cgcgggtggg cctacttcac ctatcctgcc cggctgacgc cgttggatac accaaggaaa
     6841 gtctacacga accctttggc aaaatcctgt atatcgtgcg aaaaaggatg gatataccga
     6901 aaaaatcgct ataatgaccc cgaagcaggg ttatgcagcg gaaaagcgct gcttccctgc
     6961 tgttttgtgg aatatctacc gactggaaac aggcaaatgc aggaaattac tgaactgagg
     7021 ggacaggcga gagacgatgc caaagagcta caccgacgag ctggccgagt gggttgaatc
     7081 ccgcgcggcc aagaagcgcc ggcgtgatga ggctgcggtt gcgttcctgg cggtgagggc
     7141 ggatgtcgag gcggcgttag cgtccggcta tgcgctcgtc accatttggg agcacatgcg
     7201 ggaaacgggg aaggtcaagt tctcctacga gacgttccgc tcgcacgcca ggcggcacat
     7261 caaggccaag cccgccgatg tgcccgcacc gcaggccaag gctgcggaac ccgcgccggc
     7321 acccaagacg ccggagccac ggcggccgaa gcaggggggc aaggctgaaa agccggcccc
     7381 cgctgcggcc ccgaccggct tcaccttcaa cccaacaccg gacaaaaagg atcggcgttg
     7441 ccatagtcga ttttcttcaa atagcgccga gctttgtcat caccaatttc ctttgcaaat
     7501 agctcataca cccaaacagt agaattcccc ggatccgtcg acctgcagaa aggcaggccg
     7561 ggccgtggtg gccacggcct ctaggccaga tccagcggca tctgggttag tcgagcgcgg
     7621 gccgcttccc atgtctcacc agggcgagcc tgtttcgcga tctcagcatc tgaaatcttc
     7681 ccggccttgc gcttcgctgg ggccttaccc accgccttgg cgggcttctt cggtccaaaa
     7741 ctgaacaaca gatgtgtgac cttgcgcccg gtctttcgct gcgcccactc cacctgtagc
     7801 gggctgtgct cgttgatctg cgtcacggct ggatcaagca ctcgcaactt gaagtccttg
     7861 atcgagggat accggccttc cagttgaaac cactttcgca gctggtcaat ttctatttcg
     7921 cgctggccga tgctgtccca ttgcatgagc agctcgtaaa gcctgatcgc gtgggtgctg
     7981 tccatcttgg ccacgtcagc caaggcgtat ttggtgaact gtttggtgag ttccgtcagg
     8041 tacggcagca tgtctttggt gaacctgagt tctacacggc cctcaccctc ccggtagatg
     8101 attgtttgca cccagccggt aatcatcaca ctcggtcttt tccccttgcc attgggctct
     8161 tgggttaacc ggacttcccg ccgtttcagg cgcagggccg cttctttgag ctggttgtag
     8221 gaagattcga tagggacacc cgccatcgtc gctatgtcct ccgccgtcac tgaatacatc
     8281 acttcatcgg tgacaggctc gctcctcttc acctggctaa tacaggccag aacgatccgc
     8341 tgttcctgaa cactgaggcg atacgcggcc tcgaccaggg cattgctttt gtaaaccatt
     8401 gggggtgagg ccacgttcga cattccttgt gtataagggg acactgtatc tgcgtcccac
     8461 aatacaac
//
