LOCUS       sequence-153394-        5381 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E.coli BW25113 + JBEI-12050 (pBbA5c-MevTo-BBa1002-pTrc-Mkco-aphA, 
            JBx_049165 ) + JBEI-9314 (pTrc99a-PMDsc, JBx_039366).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5381)
  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 5381)
  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: ARK2aa
FEATURES             Location/Qualifiers
     source          1..5381
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(232..237,238..254,255..261)
                     /label=trc promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    269..285
                     /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     274..296
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             613..624
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(1603..1620)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1773..1859
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1951..1978
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        1998..2089
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2090..2158,2159..2950)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPTAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2308..2327)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3121..3709
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3610..3629
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3863..3880
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3895..4035
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3961..3980)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4121..4143
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     promoter        4221..4298
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             4299..5381
                     /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
                     YIPPSTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(4318..4337)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 gcgcaacgca attaatgtga gttagcgcga attgatctgg tttgacagct tatcatcgac
       61 tgcacggtgc accaatgctt ctggcgtcag gcagccatcg gaagctgtgg tatggctgtg
      121 caggtcgtaa atcactgcat aattcgtgtc gctcaaggcg cactcccgtt ctggataatg
      181 ttttttgcgc cgacatcata acggttctgg caaatattct gaaatgagct gttgacaatt
      241 aatcatccgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca cacaggaaac
      301 agaccatgga attcaaaaga tcttttaaga aggagatata catatgaccg tttacacagc
      361 atccgttacc gcacccgtca acatcgcaac ccttaagtat tgggggaaaa gggacacgaa
      421 gttgaatctg cccaccaatt cgtccatatc agtgacttta tcgcaagatg acctcagaac
      481 gttgacctct gcggctactg cacctgagtt tgaacgcgac actttgtggt taaatggaga
      541 accacacagc atcgacaatg aaagaactca aaattgtctg cgcgacctac gccaattaag
      601 aaaggaaatg gaatcgaagg acgcctcatt gcccacatta tctcaatgga aactccacat
      661 tgtctccgaa aataactttc ctacagcagc tggtttagct tcctccgctg ctggctttgc
      721 tgcattggtc tctgcaattg ctaagttata ccaattacca cagtcaactt cagaaatatc
      781 tagaatagca agaaaggggt ctggttcagc ttgtagatcg ttgtttggcg gatacgtggc
      841 ctgggaaatg ggaaaagctg aagatggtca tgattccatg gcagtacaaa tcgcagacag
      901 ctctgactgg cctcagatga aagcttgtgt cctagttgtc agcgatatta aaaaggatgt
      961 gagttccact cagggtatgc aattgaccgt ggcaacctcc gaactattta aagaaagaat
     1021 tgaacatgtc gtaccaaaga gatttgaagt catgcgtaaa gccattgttg aaaaagattt
     1081 cgccaccttt gcaaaggaaa caatgatgga ttccaactct ttccatgcca catgtttgga
     1141 ctctttccct ccaatattct acatgaatga cacttccaag cgtatcatca gttggtgcca
     1201 caccattaat cagttttacg gagaaacaat cgttgcatac acgtttgatg caggtccaaa
     1261 tgctgtgttg tactacttag ctgaaaatga gtcgaaactc tttgcattta tctataaatt
     1321 gtttggctct gttcctggat gggacaagaa atttactact gagcagcttg aggctttcaa
     1381 ccatcaattt gaatcatcta actttactgc acgtgaattg gatcttgagt tgcaaaagga
     1441 tgttgccaga gtgattttaa ctcaagtcgg ttcaggccca caagaaacaa acgaatcttt
     1501 gattgacgca aagactggtc taccaaagga ataaggatcc aaactcgagt aactgcaggc
     1561 atgcaagctt ggctgttttg gcggatgaga gaagattttc agcctgatac agattaaatc
     1621 agaacgcaga agcggtctga taaaacagaa tttgcctggc ggcagtagcg cggtggtccc
     1681 acctgacccc atgccgaact cagaagtgaa acgccgtagc gccgatggta gtgtggggtc
     1741 tccccatgcg agagtaggga actgccaggc atcaaataaa acgaaaggct cagtcgaaag
     1801 actgggcctt tcgttttatc tgttgtttgt cggtgaacgc tctcctgagt aggacaaatc
     1861 cgccgggagc ggatttgaac gttgcgaagc aacggcccgg agggtggcgg gcaggacgcc
     1921 cgccataaac tgccaggcat caaattaagc agaaggccat cctgacggat ggcctttttg
     1981 cgtttctaca aactcttttt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     2041 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     2101 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     2161 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     2221 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     2281 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtgttgacgc
     2341 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     2401 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     2461 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     2521 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     2581 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctacagcaat
     2641 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     2701 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     2761 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     2821 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     2881 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     2941 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     3001 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     3061 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     3121 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     3181 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     3241 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
     3301 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     3361 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     3421 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     3481 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     3541 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     3601 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     3661 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     3721 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     3781 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     3841 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat
     3901 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatatggt gcactctcag
     3961 tacaatctgc tctgatgccg catagttaag ccagtataca ctccgctatc gctacgtgac
     4021 tgggtcatgg ctgcgccccg acacccgcca acacccgctg acgcgccctg acgggcttgt
     4081 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag
     4141 aggttttcac cgtcatcacc gaaacgcgcg aggcagcaga tcaattcgcg cgcgaaggcg
     4201 aagcggcatg catttacgtt gacaccatcg aatggtgcaa aacctttcgc ggtatggcat
     4261 gatagcgccc ggaagagagt caattcaggg tggtgaatgt gaaaccagta acgttatacg
     4321 atgtcgcaga gtatgccggt gtctcttatc agaccgtttc ccgcgtggtg aaccaggcca
     4381 gccacgtttc tgcgaaaacg cgggaaaaag tggaagcggc gatggcggag ctgaattaca
     4441 ttcccaaccg cgtggcacaa caactggcgg gcaaacagtc gttgctgatt ggcgttgcca
     4501 cctccagtct ggccctgcac gcgccgtcgc aaattgtcgc ggcgattaaa tctcgcgccg
     4561 atcaactggg tgccagcgtg gtggtgtcga tggtagaacg aagcggcgtc gaagcctgta
     4621 aagcggcggt gcacaatctt ctcgcgcaac gcgtcagtgg gctgatcatt aactatccgc
     4681 tggatgacca ggatgccatt gctgtggaag ctgcctgcac taatgttccg gcgttatttc
     4741 ttgatgtctc tgaccagaca cccatcaaca gtattatttt ctcccatgaa gacggtacgc
     4801 gactgggcgt ggagcatctg gtcgcattgg gtcaccagca aatcgcgctg ttagcgggcc
     4861 cattaagttc tgtctcggcg cgtctgcgtc tggctggctg gcataaatat ctcactcgca
     4921 atcaaattca gccgatagcg gaacgggaag gcgactggag tgccatgtcc ggttttcaac
     4981 aaaccatgca aatgctgaat gagggcatcg ttcccactgc gatgctggtt gccaacgatc
     5041 agatggcgct gggcgcaatg cgcgccatta ccgagtccgg gctgcgcgtt ggtgcggata
     5101 tctcggtagt gggatacgac gataccgaag acagctcatg ttatatcccg ccgtcaacca
     5161 ccatcaaaca ggattttcgc ctgctggggc aaaccagcgt ggaccgcttg ctgcaactct
     5221 ctcagggcca ggcggtgaag ggcaatcagc tgttgcccgt ctcactggtg aaaagaaaaa
     5281 ccaccctggc gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc
     5341 agctggcacg acaggtttcc cgactggaaa gcgggcagtg a
//
