LOCUS       sequence-153406-        9326 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E.coli BW25113 + JBEI-9310 (pA5c-MevT(O)-T21-MKco, JBx_039538) + 
            JBEI-9314(pTrc99a-PMDsc, JBx_039366).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9326)
  AUTHORS   Kang A, George KW, Wang G, Baidoo E, Keasling JD, Lee TS
  TITLE     Isopentenyl diphosphate (IPP)-bypass mevalonate pathways for 
            isopentenol production.
  JOURNAL   Metab Eng. 2016 Mar;34:25-35. doi: 10.1016/j.ymben.2015.12.002. Epub
            2015 Dec 17.
  PUBMED    26708516
REFERENCE   2  (bases 1 to 9326)
  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: "Metab 
            Eng."; date: "2016-03"; volume: "34:25-35. doi"; pages: " 
            10.1016/j.ymben.2015.12.002. Epub 2015 Dec 17"
COMMENT     Sequence Label: ARK2a
FEATURES             Location/Qualifiers
     source          1..9326
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(49..1131)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     1093..1112
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1132..1209)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1415..1434
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        join(1520..1525,1526..1543,1544..1550)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    1558..1574
                     /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)."
     CDS             complement(2309..2320)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             5233..5244
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     promoter        join(6048..6053,6054..6070,6071..6077)
                     /label=trc promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    6085..6101
                     /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)."
     terminator      7491..7562
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      7578..7605
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     primer_bind     complement(7633..7650)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7767..8312)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(7847..7866)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(8426..8520)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(8541..9200)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     9115..9134
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(9201..9303)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
ORIGIN
        1 gacgtcggtg cctaatgagt gagctaactt acattaattg cgttgcgctc actgcccgct
       61 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
      121 ggcggtttgc gtattgggcg ccagggtggt ttttcttttc accagtgaga cgggcaacag
      181 ctgattgccc ttcaccgcct ggccctgaga gagttgcagc aagcggtcca cgctggtttg
      241 ccccagcagg cgaaaatcct gtttgatggt ggttaacggc gggatataac atgagctgtc
      301 ttcggtatcg tcgtatccca ctaccgagat gtccgcacca acgcgcagcc cggactcggt
      361 aatggcgcgc attgcgccca gcgccatctg atcgttggca accagcatcg cagtgggaac
      421 gatgccctca ttcagcattt gcatggtttg ttgaaaaccg gacatggcac tccagtcgcc
      481 ttcccgttcc gctatcggct gaatttgatt gcgagtgaga tatttatgcc agccagccag
      541 acgcagacgc gccgagacag aacttaatgg gcccgctaac agcgcgattt gctggtgacc
      601 caatgcgacc agatgctcca cgcccagtcg cgtaccgtct tcatgggaga aaataatact
      661 gttgatgggt gtctggtcag agacatcaag aaataacgcc ggaacattag tgcaggcagc
      721 ttccacagca atggcatcct ggtcatccag cggatagtta atgatcagcc cactgacgcg
      781 ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg acgccgcttc gttctaccat
      841 cgacaccacc acgctggcac ccagttgatc ggcgcgagat ttaatcgccg cgacaatttg
      901 cgacggcgcg tgcagggcca gactggaggt ggcaacgcca atcagcaacg actgtttgcc
      961 cgccagttgt tgtgccacgc ggttgggaat gtaattcagc tccgccatcg ccgcttccac
     1021 tttttcccgc gttttcgcag aaacgtggct ggcctggttc accacgcggg aaacggtctg
     1081 ataagagaca ccggcatact ctgcgacatc gtataacgtt actggtttca cattcaccac
     1141 cctgaattga ctctcttccg ggcgctatca tgccataccg cgaaaggttt tgcgccattc
     1201 gatggtgtcc gggatctcga cgctctccct tatgcgactc ctgcattagg aagcagccca
     1261 gtagtaggtt gaggccgttg agcaccgccg ccgcaaggaa tggtgcatgc aaggagatgg
     1321 cgcccaacag tcccccggcc acggggcctg ccaccatacc cacgccgaaa caagcgctca
     1381 tgagcccgaa gtggcgagcc cgatcttccc catcggtgat gtcggcgata taggcgccag
     1441 caaccgcacc tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag aggatcgaga
     1501 tcgtttaggc accccaggct ttacacttta tgcttccggc tcgtataatg tgtggaattg
     1561 tgagcggata acaatttcag aattcaaaag atcttaggag gaatataaaa tgaaaaattg
     1621 tgtcatcgtc agtgcggtac gtactgctat cggtagtttt aacggttcac tcgcttccac
     1681 cagcgccatc gacctggggg cgacagtaat taaagccgcc attgaacgtg caaaaatcga
     1741 ttcacaacac gttgatgaag tgattatggg taacgtgtta caagccgggc tggggcaaaa
     1801 tccggcgcgt caggcactgt taaaaagcgg gctggcagaa acggtgtgcg gattcacggt
     1861 caataaagta tgtggttcgg gtcttaaaag tgtggcgctt gccgcccagg ccattcaggc
     1921 aggtcaggcg cagagcattg tggcgggggg tatggaaaat atgagtttag ccccctactt
     1981 actcgatgca aaagcacgct ctggttatcg tcttggagac ggacaggttt atgacgtaat
     2041 cctgcgcgat ggcctgatgt gcgccaccca tggttatcat atggggatta ccgccgaaaa
     2101 cgtggctaaa gagtacggaa ttacccgtga aatgcaggat gaactggcgc tacattcaca
     2161 gcgtaaagcg gcagccgcaa ttgagtccgg tgcttttaca gccgaaatcg tcccggtaaa
     2221 tgttgtcact cgaaagaaaa ccttcgtctt cagtcaagac gagttcccga aagcgaactc
     2281 aacggctgaa gcgttaggtg cattgcgccc ggccttcgat aaagcaggaa cagtcaccgc
     2341 tgggaacgcg tctggtatta acgacggtgc tgccgctctg gtgattatgg aagaatctgc
     2401 ggcgctggca gcaggcctta cccccctggc tcgcattaaa agttatgcca gcggtggcgt
     2461 gccccccgca ttgatgggta tggggccagt acctgccacg caaaaagcgt tacaactggc
     2521 ggggctgcaa ctggcggata ttgatctcat tgaggctaat gaagcatttg ctgcacagtt
     2581 ccttgccgtt gggaaaaacc tgggctttga ttctgagaaa gtgaatgtca acggcggggc
     2641 catcgcgctc gggcatccta tcggtgccag tggtgctcgt attctggtca cactattaca
     2701 tgccatgcag gcacgcgata aaacgctggg gctggcaaca ctgtgcattg gcggcggtca
     2761 gggaattgcg atggtgattg aacggttgaa ttgaggatct tgaattaagg aggacagcta
     2821 aatgaaactc tcaactaaac tttgttggtg tggtattaaa ggaagactta ggccgcaaaa
     2881 gcaacaacaa ttacacaata caaacttgca aatgactgaa ctaaaaaaac aaaagaccgc
     2941 tgaacaaaaa accagacctc aaaatgtcgg tattaaaggt atccaaattt acatcccaac
     3001 tcaatgtgtc aaccaatctg agctagagaa atttgatggc gtttctcaag gtaaatacac
     3061 aattggtctg ggccaaacca acatgtcttt tgtcaatgac agagaagata tctactcgat
     3121 gtccctaact gttttgtcta agttgatcaa gagttacaac atcgacacca acaaaattgg
     3181 tagattagaa gtcggtactg aaactctgat tgacaagtcc aagtctgtca agtctgtctt
     3241 gatgcaattg tttggtgaaa acactgacgt cgaaggtatt gacacgctta atgcctgtta
     3301 cggtggtacc aacgcgttgt tcaactcttt gaactggatt gaatctaacg catgggatgg
     3361 tagagacgcc attgtagttt gcggtgatat tgccatctac gataagggtg ccgcaagacc
     3421 aaccggtggt gccggtactg ttgctatgtg gatcggtcct gatgctccaa ttgtatttga
     3481 ctctgtaaga gcttcttaca tggaacacgc ctacgatttt tacaagccag atttcaccag
     3541 cgaatatcct tacgtcgatg gtcatttttc attaacttgt tacgtcaagg ctcttgatca
     3601 agtttacaag agttattcca agaaggctat ttctaaaggg ttggttagcg atcccgctgg
     3661 ttcggatgct ttgaacgttt tgaaatattt cgactacaac gttttccatg ttccaacctg
     3721 taaattggtc acaaaatcat acggtagatt actatataac gatttcagag ccaatcctca
     3781 attgttccca gaagttgacg ccgaattagc tactcgcgat tatgacgaat ctttaaccga
     3841 taagaacatt gaaaaaactt ttgttaatgt tgctaagcca ttccacaaag agagagttgc
     3901 ccaatctttg attgttccaa caaacacagg taacatgtac accgcatctg tttatgccgc
     3961 ctttgcatct ctattaaact atgttggatc tgacgactta caaggcaagc gtgttggttt
     4021 attttcttac ggttccggtt tagctgcatc tctatattct tgcaaaattg ttggtgacgt
     4081 ccaacatatt atcaaggaat tagatattac taacaaatta gccaagagaa tcaccgaaac
     4141 tccaaaggat tacgaagctg ccatcgaatt gagagaaaat gcccatttga agaagaactt
     4201 caaacctcaa ggttccattg agcatttgca aagtggtgtt tactacttga ccaacatcga
     4261 tgacaaattt agaagatcct acgatgttaa aaaatgagga tctaaaataa ggaggattac
     4321 actatggttt taaccaataa aacagtcatt tctggatcga aagtcaaaag tttatcatct
     4381 gcgcaatcga gctcatcagg accttcatca tctagtgagg aagatgattc ccgcgatatt
     4441 gaaagcttgg ataagaaaat acgtccttta gaagaattag aagcattatt aagtagtgga
     4501 aatacaaaac aattgaagaa caaagaggtc gctgccttgg ttattcacgg taagttacct
     4561 ttgtacgctt tggagaaaaa attaggtgat actacgagag cggttgcggt acgtaggaag
     4621 gctctttcaa ttttggcaga agctcctgta ttagcatctg atcgtttacc atataaaaat
     4681 tatgactacg accgcgtatt tggcgcttgt tgtgaaaatg ttataggtta catgcctttg
     4741 cccgttggtg ttataggccc cttggttatc gatggtacat cttatcatat accaatggca
     4801 actacagagg gttgtttggt agcttctgcc atgcgtggct gtaaggcaat caatgctggc
     4861 ggtggtgcaa caactgtttt aactaaggat ggtatgacaa gaggcccagt agtccgtttc
     4921 ccaactttga aaagatccgg tgcctgtaag atatggttag actcagaaga gggacaaaac
     4981 gcaattaaaa aagcttttaa ctctacatca agatttgcac gtctgcaaca tattcaaact
     5041 tgtctagcag gagatttact cttcatgaga tttagaacaa ctactggtga cgcaatgggt
     5101 atgaatatga tttctaaagg tgtcgaatac tcattaaagc aaatggtaga agagtatggc
     5161 tgggaagata tggaggttgt ctccgtttct ggtaactact gtaccgacaa aaaaccagct
     5221 gccatcaact ggatcgaagg tcgtggtaag agtgtcgtcg cagaagctac tattcctggt
     5281 gatgttgtca gaaaagtgtt aaaaagtgat gtttccgcat tggttgagtt gaacattgct
     5341 aagaatttgg ttggatctgc aatggctggg tctgttggtg gatttaacgc acatgcagct
     5401 aatttagtga cagctgtttt cttggcatta ggacaagatc ctgcacaaaa tgttgaaagt
     5461 tccaactgta taacattgat gaaagaagtg gacggtgatt tgagaatttc cgtatccatg
     5521 ccatccatcg aagtaggtac catcggtggt ggtactgttc tagaaccaca aggtgccatg
     5581 ttggacttat taggtgtaag aggcccgcat gctaccgctc ctggtaccaa cgcacgtcaa
     5641 ttagcaagaa tagttgcctg tgccgtcttg gcaggtgaat tatccttatg tgctgcccta
     5701 gcagccggcc atttggttca aagtcatatg acccacaaca ggaaacctgc tgaaccaaca
     5761 aaacctaaca atttggacgc cactgatata aatcgtttga aagatgggtc cgtcacctgc
     5821 attaaatcct gaggatctcg caaaaaaccc cgcttcggcg gggttttttc gccgactgca
     5881 cggtgcacca atgcttctgg cgtcaggcag ccatcggaag ctgtggtatg gctgtgcagg
     5941 tcgtaaatca ctgcataatt cgtgtcgctc aaggcgcact cccgttctgg ataatgtttt
     6001 ttgcgccgac atcataacgg ttctggcaaa tattctgaaa tgagctgttg acaattaatc
     6061 atccggctcg tataatgtgt ggaattgtga gcggataaca atttcaggat ctaggaggaa
     6121 ataaccatgt ctctgccatt cctgacgtct gcgccaggta aggtgatcat cttcggcgag
     6181 cactctgcgg tgtacaataa gccggccgtc gccgcctctg tgtctgcgtt acgcacctac
     6241 ctgctgatca gcgaatcttc tgcaccggac acgatcgagc tggactttcc ggacatcagc
     6301 ttcaaccaca agtggagcat caacgacttc aacgcgatca cggaggacca ggtgaacagc
     6361 caaaagctgg ccaaagccca gcaagcaacc gacggtctgt ctcaggagct ggtgtctctg
     6421 ctggacccgc tgttagcgca gttaagcgag agcttccatt accacgccgc gttctgcttc
     6481 ctgtacatgt tcgtttgcct gtgcccgcac gcaaagaaca tcaagttcag cctgaagagc
     6541 acgctgccga ttggcgcagg cttaggctct agcgcatcta tcagcgtgag cctggcgctg
     6601 gcgatggcct atctgggtgg cctgattggc agcaacgacc tggagaaact gagcgaaaac
     6661 gacaagcaca tcgtgaacca gtgggccttt atcggcgaga agtgcattca tggcaccccg
     6721 agcggcattg acaacgcagt tgccacgtat ggcaacgccc tgctgttcga gaaagacagc
     6781 cacaacggca cgatcaacac gaacaacttc aagttcctgg acgacttccc ggcgatcccg
     6841 atgattctga cctacacccg tatcccacgc agcaccaagg atttagtcgc ccgcgtgcgt
     6901 gttttagtca ccgaaaagtt cccggaggtg atgaagccga tcctggacgc gatgggcgag
     6961 tgcgcgctgc agggtctgga gatcatgacc aagctgagca agtgcaaggg caccgacgat
     7021 gaggcggtgg agaccaacaa tgagctgtac gagcagctgc tggagctgat ccgtatcaat
     7081 cacggcctgc tggtctctat cggtgtgtct cacccgggcc tggaactgat caaaaacctg
     7141 agcgacgacc tgcgcattgg ctctacgaaa ttaacgggtg caggtggcgg tggctgctct
     7201 ttaacgctgc tgcgccgtga cattacgcag gagcaaatcg acagcttcaa gaagaagctg
     7261 caggacgact tcagctacga gacgttcgag acggacctgg gcggcacggg ctgttgcctg
     7321 ctgagcgcca aaaatctgaa caaggacctg aagatcaaaa gcctggtgtt ccagctgttc
     7381 gaaaacaaga cgaccacgaa gcagcagatc gacgacctgt tactgccggg taacaccaat
     7441 ctgccgtgga cgtcttaagg atccaaactc gagtaaggat ctccaggcat caaataaaac
     7501 gaaaggctca gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg gtgaacgctc
     7561 tctactagag tcacactggc tcaccttcgg gtgggccttt ctgcgtttat acctagggat
     7621 atattccgct tcctcgctca ctgactcgct acgctcggtc gttcgactgc ggcgagcgga
     7681 aatggcttac gaacggggcg gagatttcct ggaagatgcc aggaagatac ttaacaggga
     7741 agtgagaggg ccgcggcaaa gccgtttttc cataggctcc gcccccctga caagcatcac
     7801 gaaatctgac gctcaaatca gtggtggcga aacccgacag gactataaag ataccaggcg
     7861 tttccccctg gcggctccct cgtgcgctct cctgttcctg cctttcggtt taccggtgtc
     7921 attccgctgt tatggccgcg tttgtctcat tccacgcctg acactcagtt ccgggtaggc
     7981 agttcgctcc aagctggact gtatgcacga accccccgtt cagtccgacc gctgcgcctt
     8041 atccggtaac tatcgtcttg agtccaaccc ggaaagacat gcaaaagcac cactggcagc
     8101 agccactggt aattgattta gaggagttag tcttgaagtc atgcgccggt taaggctaaa
     8161 ctgaaaggac aagttttggt gactgcgctc ctccaagcca gttacctcgg ttcaaagagt
     8221 tggtagctca gagaaccttc gaaaaaccgc cctgcaaggc ggttttttcg ttttcagagc
     8281 aagagattac gcgcagacca aaacgatctc aagaagatca tcttattaat cagataaaat
     8341 atttctagat ttcagtgcaa tttatctctt caaatgtagc acctgaagtc agccccatac
     8401 gatataagtt gttactagtg cttggattct caccaataaa aaacgcccgg cggcaaccga
     8461 gcgttctgaa caaatccaga tggagttctg aggtcattac tggatctatc aacaggagtc
     8521 caagcgagct cgatatcaaa ttacgccccg ccctgccact catcgcagta ctgttgtaat
     8581 tcattaagca ttctgccgac atggaagcca tcacaaacgg catgatgaac ctgaatcgcc
     8641 agcggcatca gcaccttgtc gccttgcgta taatatttgc ccatggtgaa aacgggggcg
     8701 aagaagttgt ccatattggc cacgtttaaa tcaaaactgg tgaaactcac ccagggattg
     8761 gctgagacga aaaacatatt ctcaataaac cctttaggga aataggccag gttttcaccg
     8821 taacacgcca catcttgcga atatatgtgt agaaactgcc ggaaatcgtc gtggtattca
     8881 ctccagagcg atgaaaacgt ttcagtttgc tcatggaaaa cggtgtaaca agggtgaaca
     8941 ctatcccata tcaccagctc accgtctttc attgccatac gaaattccgg atgagcattc
     9001 atcaggcggg caagaatgtg aataaaggcc ggataaaact tgtgcttatt tttctttacg
     9061 gtctttaaaa aggccgtaat atccagctga acggtctggt tataggtaca ttgagcaact
     9121 gactgaaatg cctcaaaatg ttctttacga tgccattggg atatatcaac ggtggtatat
     9181 ccagtgattt ttttctccat tttagcttcc ttagctcctg aaaatctcga taactcaaaa
     9241 aatacgcccg gtagtgatct tatttcatta tggtgaaagt tggaacctct tacgtgccga
     9301 tcaacgtctc attttcgcca gatatc
//
