LOCUS       sequence-94224-e        8821 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pKT230 derivative; PBAD_rfp; cmR.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8821)
  AUTHORS   Bi C, Su P, Muller J, Yeh YC, Chhabra SR, Beller HR, Singer SW, 
            Hillson NJ
  TITLE     Development of a broad-host synthetic biology toolbox for Ralstonia 
            eutropha and its application to engineering hydrocarbon biofuel 
            production.
  JOURNAL   Microb Cell Fact. 2013 Nov 13;12:107. doi: 10.1186/1475-2859-12-107.
  PUBMED    24219429
REFERENCE   2  (bases 1 to 8821)
  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: "Microb Cell
            Fact."; date: "2013-11-13"; volume: "12:107. doi"; pages: " 
            10.1186/1475-2859-12-107"
COMMENT     Sequence Label: pKTrfp
FEATURES             Location/Qualifiers
     source          1..8821
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      396..790
                     /label=RSF1010 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     RSF1010; requires the RSF1010 RepA/B/C proteins for 
                     replication (Scholz et al., 1989)"
     oriT            1132..1219
                     /label=RSF1010 oriT
                     /note="origin of transfer of the broad-host-range plasmid 
                     RSF1010 (Scholz et al., 1989)"
     CDS             2458..3429
                     /codon_start=1
                     /gene="repB"
                     /product="replication protein B of the broad-host-range 
                     plasmid RSF1010 (Scholz et al., 1989)"
                     /label=RSF1010 RepB
                     /translation="MKNDRTLQAIGRQLKAMGCERFDIGVRDATTGQMMNREWSAAEVL
                     QNTPWLKRMNAQGNDVYIRPAEQERHGLVLVDDLSEFDLDDMKAEGREPALVVETSPKN
                     YQAWVKVADAAGGELRGQIARTLASEYDADPASADSRHYGRLAGFTNRKDKHTTRAGYQ
                     PWVLLRESKGKTATAGPALVQQAGQQIEQAQRQQEKARRLASLELPERQLSRHRRTALD
                     EYRSEMAGLVKRFGDDLSKCDFIAAQKLASRGRSAEEIGKAMAEASPALAERKPGHEAD
                     YIERTVSKVMGLPSVQLARAELARAPAPRQRGMDRGGPDFSM"
     CDS             3940..4779
                     /codon_start=1
                     /gene="repA"
                     /product="replication protein A of the broad-host-range 
                     plasmid RSF1010 (Scholz et al., 1989)"
                     /label=RSF1010 RepA
                     /translation="MATHKPINILEAFAAAPPPLDYVLPNMVAGTVGALVSPGGAGKSM
                     LALQLAAQIAGGPDLLEVGELPTGPVIYLPAEDPPTAIHHRLHALGAHLSAEERQAVAD
                     GLLIQPLIGSLPNIMAPEWFDGLKRAAEGRRLMVLDTLRRFHIEEENASGPMAQVIGRM
                     EAIAADTGCSIVFLHHASKGAAMMGAGDQQQASRGSSVLVDNIRWQSYLSSMTSAEAEE
                     WGVDDDQRRFFVRFGVSKANYGAPFADRWFRRHDGGVLKPAVLERQRKSKGVPRGEA"
     CDS             4766..5617
                     /codon_start=1
                     /gene="repC"
                     /product="replication protein C of the broad-host-range 
                     plasmid RSF1010 (Scholz et al., 1989)"
                     /label=RSF1010 RepC
                     /translation="MVKPKNKHSLSHVRHDPAHCLAPGLFRALKRGERKRSKLDVTYDY
                     GDGKRIEFSGPEPLGADDLRILQGLVAMAGPNGLVLGPEPKTEGGRQLRLFLEPKWEAV
                     TAECHVVKGSYRALAKEIGAEVDSGGALKHIQDCIERLWKVSIIAQNGRKRQGFRLLSE
                     YASDEADGRLYVALNPLIAQAVMGGGQHVRISMDEVRALDSETARLLHQRLCGWIDPGK
                     TGKASIDTLCGYVWPSEASGSTMRKRRQRVREALPELVALGWTVTEFAAGKYDITRPKA
                     AG"
     terminator      complement(5836..5930)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(5951..6610)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     6525..6544
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(6611..6713)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             complement(6743..7621)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        7648..7932
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     7855..7874
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             7982..8659
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     terminator      8692..8763
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      8779..8806
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
ORIGIN
        1 ctgaaagcga ccaggtgctc ggcgtggcaa gactcgcagc gaacccgtag aaagccatgc
       61 tccagccgcc cgcattggag aaattcttca aattcccgtt gcacatagcc cggcaattcc
      121 tttccctgct ctgccataag cgcagcgaat gccgggtaat actcgtcaac gatctgatag
      181 agaagggttt gctcgggtcg gtggctctgg taacgaccag tatcccgatc ccggctggcc
      241 gtcctggccg ccacatgagg catgttccgc gtccttgcaa tactgtgttt acatacagtc
      301 tatcgcttag cggaaagttc ttttaccctc agccgaaatg cctgccgttg ctagacattg
      361 ccagccagtg cccgtcactc ccgtactaac tgtcacgaac ccctgcaata actgtcacgc
      421 ccccctgcaa taactgtcac gaacccctgc aataactgtc acgcccccaa acctgcaaac
      481 ccagcagggg cgggggctgg cggggtgttg gaaaaatcca tccatgatta tctaagaata
      541 atccactagg cgcggttatc agcgcccttg tggggcgctg ctgcccttgc ccaatatgcc
      601 cggccagagg ccggatagct ggtctattcg ctgcgctagg ctacacaccg ccccaccgct
      661 gcgcggcagg gggaaaggcg ggcaaagccc gctaaacccc acaccaaacc ccgcagaaat
      721 acgctggagc gcttttagcc gctttagcgg cctttccccc tacccgaagg gtgggggcgc
      781 gtgtgcagcc ccgcagggcc tgtctcggtc gatcattcag cccggctcat ccttctggcg
      841 tggcggcaga ccgaacaagg cgcggtcgtg gtcgcgttca aggtacgcat ccattgccgc
      901 catgagccga tcctccggcc actcgctgct gttcaccttg gccaaaatca tggcccccac
      961 cagcaccttg cgccttgttt cgttcttgcg ctcttgctgc tgttcccttg cccgcacccg
     1021 ctgaatttcg gcattgattc gcgctcgttg ttcttcgagc ttggccagcc gatccgccgc
     1081 cttgttgctc cccttaacca tcttgacacc ccattgttaa tgtgctgtct cgtaggctat
     1141 catggaggca cagcggcggc aatcccgacc ctactttgta ggggagggcg cacttaccgg
     1201 tttctcttcg agaaactggc ctaacggcca cccttcgggc ggtgcgctct ccgagggcca
     1261 ttgcatggag ccgaaaagca aaagcaacag cgaggcagca tggcgattta tcaccttacg
     1321 gcgaaaaccg gcagcaggtc gggcggccaa tcggccaggg ccaaggccga ctacatccag
     1381 cgcgaaggca agtatgcccg cgacatggat gaagtcttgc acgccgaatc cgggcacatg
     1441 ccggagttcg tcgagcggcc cgccgactac tgggatgctg ccgacctgta tgaacgcgcc
     1501 aatgggcggc tgttcaagga ggtcgaattt gccctgccgg tcgagctgac cctcgaccag
     1561 cagaaggcgc tggcgtccga gttcgcccag cacctgaccg gtgccgagcg cctgccgtat
     1621 acgctggcca tccatgccgg tggcggcgag aacccgcact gccacctgat gatctccgag
     1681 cggatcaatg acggcatcga gcggcccgcc gctcagtggt tcaagcggta caacggcaag
     1741 accccggaga agggcggggc acagaagacc gaagcgctca agcccaaggc atggcttgag
     1801 cagacccgcg aggcatgggc cgaccatgcc aaccgggcat tagagcgggc tggccacgac
     1861 gcccgcattg accacagaac acttgaggcg cagggcatcg agcgcctgcc cggtgttcac
     1921 ctggggccga acgtggtgga gatggaaggc cggggcatcc gcaccgaccg ggcagacgtg
     1981 gccctgaaca tcgacaccgc caacgcccag atcatcgact tacaggaata ccgggaggca
     2041 atagaccatg aacgcaatcg acagagtgaa gaaatccaga ggcatcaacg agttagcgga
     2101 gcagatcgaa ccgctggccc agagcatggc gacactggcc gacgaagccc ggcaggtcat
     2161 gagccagacc cagcaggcca gcgaggcgca ggcggcggag tggctgaaag cccagcgcca
     2221 gacaggggcg gcatgggtgg agctggccaa agagttgcgg gaggtagccg ccgaggtgag
     2281 cagcgccgcg cagagcgccc ggagcgcgtc gcgggggtgg cactggaagc tatggctaac
     2341 cgtgatgctg gcttccatga tgcctacggt ggtgctgctg atcgcatcgt tgctcttgct
     2401 cgacctgacg ccactgacaa ccgaggacgg ctcgatctgg ctgcgcttgg tggcccgatg
     2461 aagaacgaca ggactttgca ggccataggc cgacagctca aggccatggg ctgtgagcgc
     2521 ttcgatatcg gcgtcaggga cgccaccacc ggccagatga tgaaccggga atggtcagcc
     2581 gccgaagtgc tccagaacac gccatggctc aagcggatga atgcccaggg caatgacgtg
     2641 tatatcaggc ccgccgagca ggagcggcat ggtctggtgc tggtggacga cctcagcgag
     2701 tttgacctgg atgacatgaa agccgagggc cgggagcctg ccctggtagt ggaaaccagc
     2761 ccgaagaact atcaggcatg ggtcaaggtg gccgacgccg caggcggtga acttcggggg
     2821 cagattgccc ggacgctggc cagcgagtac gacgccgacc cggccagcgc cgacagccgc
     2881 cactatggcc gcttggcggg cttcaccaac cgcaaggaca agcacaccac ccgcgccggt
     2941 tatcagccgt gggtgctgct gcgtgaatcc aagggcaaga ccgccaccgc tggcccggcg
     3001 ctggtgcagc aggctggcca gcagatcgag caggcccagc ggcagcagga gaaggcccgc
     3061 aggctggcca gcctcgaact gcccgagcgg cagcttagcc gccaccggcg cacggcgctg
     3121 gacgagtacc gcagcgagat ggccgggctg gtcaagcgct tcggtgatga cctcagcaag
     3181 tgcgacttta tcgccgcgca gaagctggcc agccggggcc gcagtgccga ggaaatcggc
     3241 aaggccatgg ccgaggccag cccagcgctg gcagagcgca agcccggcca cgaagcggat
     3301 tacatcgagc gcaccgtcag caaggtcatg ggtctgccca gcgtccagct tgcgcgggcc
     3361 gagctggcac gggcaccggc accccgccag cgaggcatgg acaggggcgg gccagatttc
     3421 agcatgtagt gcttgcgttg gtactcacgc ctgttatact atgagtactc acgcacagaa
     3481 gggggtttta tggaatacga aaaaagcgct tcagggtcgg tctacctgat caaaagtgac
     3541 aagggctatt ggttgcccgg tggctttggt tatacgtcaa acaaggccga ggctggccgc
     3601 ttttcagtcg ctgatatggc cagccttaac cttgacggct gcaccttgtc cttgttccgc
     3661 gaagacaagc ctttcggccc cggcaagttt ctcggtgact gatatgaaag accaaaagga
     3721 caagcagacc ggcgacctgc tggccagccc tgacgctgta cgccaagcgc gatatgccga
     3781 gcgcatgaag gccaaaggga tgcgtcagcg caagttctgg ctgaccgacg acgaatacga
     3841 ggcgctgcgc gagtgcctgg aagaactcag agcggcgcag ggcgggggta gtgaccccgc
     3901 cagcgcctaa ccaccaactg cctgcaaagg aggcaatcaa tggctaccca taagcctatc
     3961 aatattctgg aggcgttcgc agcagcgccg ccaccgctgg actacgtttt gcccaacatg
     4021 gtggccggta cggtcggggc gctggtgtcg cccggtggtg ccggtaaatc catgctggcc
     4081 ctgcaactgg ccgcacagat tgcaggcggg ccggatctgc tggaggtggg cgaactgccc
     4141 accggcccgg tgatctacct gcccgccgaa gacccgccca ccgccattca tcaccgcctg
     4201 cacgcccttg gggcgcacct cagcgccgag gaacggcaag ccgtggctga cggcctgctg
     4261 atccagccgc tgatcggcag cctgcccaac atcatggccc cggagtggtt cgacggcctc
     4321 aagcgcgccg ccgagggccg ccgcctgatg gtgctggaca cgctgcgccg gttccacatc
     4381 gaggaagaaa acgccagcgg ccccatggcc caggtcatcg gtcgcatgga ggccatcgcc
     4441 gccgataccg ggtgctctat cgtgttcctg caccatgcca gcaagggcgc ggccatgatg
     4501 ggcgcaggcg accagcagca ggccagccgg ggcagctcgg tactggtcga taacatccgc
     4561 tggcagtcct acctgtcgag catgaccagc gccgaggccg aggaatgggg tgtggacgac
     4621 gaccagcgcc ggttcttcgt ccgcttcggt gtgagcaagg ccaactatgg cgcaccgttc
     4681 gctgatcggt ggttcaggcg gcatgacggc ggggtgctca agcccgccgt gctggagagg
     4741 cagcgcaaga gcaagggggt gccccgtggt gaagcctaag aacaagcaca gcctcagcca
     4801 cgtccggcac gacccggcgc actgtctggc ccccggcctg ttccgtgccc tcaagcgggg
     4861 cgagcgcaag cgcagcaagc tggacgtgac gtatgactac ggcgacggca agcggatcga
     4921 gttcagcggc ccggagccgc tgggcgctga tgatctgcgc atcctgcaag ggctggtggc
     4981 catggctggg cctaatggcc tagtgcttgg cccggaaccc aagaccgaag gcggacggca
     5041 gctccggctg ttcctggaac ccaagtggga ggccgtcacc gctgaatgcc atgtggtcaa
     5101 aggtagctat cgggcgctgg caaaggaaat cggggcagag gtcgatagtg gtggggcgct
     5161 caagcacata caggactgca tcgagcgcct ttggaaggta tccatcatcg cccagaatgg
     5221 ccgcaagcgg caggggtttc ggctgctgtc ggagtacgcc agcgacgagg cggacgggcg
     5281 cctgtacgtg gccctgaacc ccttgatcgc gcaggccgtc atgggtggcg gccagcatgt
     5341 gcgcatcagc atggacgagg tgcgggcgct ggacagcgaa accgcccgcc tgctgcacca
     5401 gcggctgtgt ggctggatcg accccggcaa aaccggcaag gcttccatag ataccttgtg
     5461 cggctatgtc tggccgtcag aggccagtgg ttcgaccatg cgcaagcgcc gccagcgggt
     5521 gcgcgaggcg ttgccggagc tggtcgcgct gggctggacg gtaaccgagt tcgcggcggg
     5581 caagtacgac atcacccggc ccaaggcggc aggctgaccc cccccactct attgtaaaca
     5641 agacattttt atcttttata ttcaatggct tattttcctg ctaattggta ataccatgaa
     5701 aaataccatg ctcagaaaag gcttaacaat attttgaaaa attgcctact gagcgctgcc
     5761 gcacagctcc ataggccgct ttcctggctt tgcttccaga tgtatgctct tctgctcctg
     5821 catactagtg cttggattct caccaataaa aaacgcccgg cggcaaccga gcgttctgaa
     5881 caaatccaga tggagttctg aggtcattac tggatctatc aacaggagtc caagcgagct
     5941 cgatatcaaa ttacgccccg ccctgccact catcgcagta ctgttgtaat tcattaagca
     6001 ttctgccgac atggaagcca tcacaaacgg catgatgaac ctgaatcgcc agcggcatca
     6061 gcaccttgtc gccttgcgta taatatttgc ccatggtgaa aacgggggcg aagaagttgt
     6121 ccatattggc cacgtttaaa tcaaaactgg tgaaactcac ccagggattg gctgagacga
     6181 aaaacatatt ctcaataaac cctttaggga aataggccag gttttcaccg taacacgcca
     6241 catcttgcga atatatgtgt agaaactgcc ggaaatcgtc gtggtattca ctccagagcg
     6301 atgaaaacgt ttcagtttgc tcatggaaaa cggtgtaaca agggtgaaca ctatcccata
     6361 tcaccagctc accgtctttc attgccatac gaaattccgg atgagcattc atcaggcggg
     6421 caagaatgtg aataaaggcc ggataaaact tgtgcttatt tttctttacg gtctttaaaa
     6481 aggccgtaat atccagctga acggtctggt tataggtaca ttgagcaact gactgaaatg
     6541 cctcaaaatg ttctttacga tgccattggg atatatcaac ggtggtatat ccagtgattt
     6601 ttttctccat tttagcttcc ttagctcctg aaaatctcga taactcaaaa aatacgcccg
     6661 gtagtgatct tatttcatta tggtgaaagt tggaacctct tacgtgccga tcaacgtctc
     6721 attttcgcca gatatcgacg tcttatgaca acttgacggc tacatcattc actttttctt
     6781 cacaaccggc acggaactcg ctcgggctgg ccccggtgca ttttttaaat acccgcgaga
     6841 aatagagttg atcgtcaaaa ccaacattgc gaccgacggt ggcgataggc atccgggtgg
     6901 tgctcaaaag cagcttcgcc tggctgatac gttggtcctc gcgccagctt aagacgctaa
     6961 tccctaactg ctggcggaaa agatgtgaca gacgcgacgg cgacaagcaa acatgctgtg
     7021 cgacgctggc gatatcaaaa ttgctgtctg ccaggtgatc gctgatgtac tgacaagcct
     7081 cgcgtacccg attatccatc ggtggatgga gcgactcgtt aatcgcttcc atgcgccgca
     7141 gtaacaattg ctcaagcaga tttatcgcca gcagctccga atagcgccct tccccttgcc
     7201 cggcgttaat gatttgccca aacaggtcgc tgaaatgcgg ctggtgcgct tcatccgggc
     7261 gaaagaaccc cgtattggca aatattgacg gccagttaag ccattcatgc cagtaggcgc
     7321 gcggacgaaa gtaaacccac tggtgatacc attcgcgagc ctccggatga cgaccgtagt
     7381 gatgaatctc tcctggcggg aacagcaaaa tatcacccgg tcggcaaaca aattctcgtc
     7441 cctgattttt caccaccccc tgaccgcgaa tggtgagatt gagaatataa cctttcattc
     7501 ccagcggtcg gtcgataaaa aaatcgagat aaccgttggc ctcaatcggc gttaaacccg
     7561 ccaccagatg ggcattaaac gagtatcccg gcagcagggg atcattttgc gcttcagcca
     7621 tacttttcat actcccgcca ttcagagaag aaaccaattg tccatattgc atcagacatt
     7681 gccgtcactg cgtcttttac tggctcttct cgctaaccaa accggtaacc ccgcttatta
     7741 aaagcattct gtaacaaagc gggaccaaag ccatgacaaa aacgcgtaac aaaagtgtct
     7801 ataatcacgg cagaaaagtc cacattgatt atttgcacgg cgtcacactt tgctatgcca
     7861 tagcattttt atccataaga ttagcggatt ctacctgacg ctttttatcg caactctcta
     7921 ctgtttctcc atacccgttt ttttgggaat tcaaaagatc ttttaagaag gagatataca
     7981 tatggcgagt agcgaagacg ttatcaaaga gttcatgcgt ttcaaagttc gtatggaagg
     8041 ttccgttaac ggtcacgagt tcgaaatcga aggtgaaggt gaaggtcgtc cgtacgaagg
     8101 tacccagacc gctaaactga aagttaccaa aggtggtccg ctgccgttcg cttgggacat
     8161 cctgtccccg cagttccagt acggttccaa agcttacgtt aaacacccgg ctgacatccc
     8221 ggactacctg aaactgtcct tcccggaagg tttcaaatgg gaacgtgtta tgaacttcga
     8281 agacggtggt gttgttaccg ttacccagga ctcctccctg caagacggtg agttcatcta
     8341 caaagttaaa ctgcgtggta ccaacttccc gtccgacggt ccggttatgc agaaaaaaac
     8401 catgggttgg gaagcttcca ccgaacgtat gtacccggaa gacggtgctc tgaaaggtga
     8461 aatcaaaatg cgtctgaaac tgaaagacgg tggtcactac gacgctgaag ttaaaaccac
     8521 ctacatggct aaaaaaccgg ttcagctgcc gggtgcttac aaaaccgaca tcaaactgga
     8581 catcacctcc cacaacgaag actacaccat cgttgaacag tacgaacgtg ctgaaggtcg
     8641 tcactccacc ggtgcttaag gatccaaact cgagtaagga tctccaggca tcaaataaaa
     8701 cgaaaggctc agtcgaaaga ctgggccttt cgttttatct gttgtttgtc ggtgaacgct
     8761 ctctactaga gtcacactgg ctcaccttcg ggtgggcctt tctgcgttta tacctagggc
     8821 c
//
