LOCUS       sequence-212816-        6708 bp    DNA     circular SYN 15-APR-2025
DEFINITION  SaGGR site-directed mutation of I206F.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6708)
  AUTHORS   Kung Y, McAndrew RP, Xie X, Liu CC, Pereira JH, Adams PD, Keasling 
            JD
  TITLE     Constructing tailored isoprenoid products by structure-guided 
            modification of geranylgeranyl reductase.
  JOURNAL   Structure. 2014 Jul 8;22(7):1028-36. doi: 10.1016/j.str.2014.05.007.
            Epub 2014 Jun 19.
  PUBMED    24954619
REFERENCE   2  (bases 1 to 6708)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.str.2014.05.007"; journalName: "Structure"; date: 
            "2014-07-8- 8"; volume: "22"; issue: "7"; pages: "1028-36"
COMMENT     Sequence Label: pSKB3SaGGR I206F (JBEI-3678)
FEATURES             Location/Qualifiers
     source          1..6708
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     907..926
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(946..1023)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1229..1248
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1332..1351
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1332..1350
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1351..1375
                     /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)."
     RBS             1390..1412
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1431..1448
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1470..1490
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2904..2921
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2974..2992)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2988..3035
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3072..3527
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3159..3178)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3369..3390
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3619..4434)
                     /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(3619..4434)
                     /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     4342..4361
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4426..4445
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4556..5144
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5045..5064
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5298..5315
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5330..5469
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5395..5414)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5555..5577
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5571..5762)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(6571..6708,1..945))
                     /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"
ORIGIN
        1 gcccttcacc gcctggccct gagagagttg cagcaagcgg tccacgctgg tttgccccag
       61 caggcgaaaa tcctgtttga tggtggttaa cggcgggata taacatgagc tgtcttcggt
      121 atcgtcgtat cccactaccg agatatccgc accaacgcgc agcccggact cggtaatggc
      181 gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg gaacgatgcc
      241 ctcattcagc atttgcatgg tttgttgaaa accggacatg gcactccagt cgccttcccg
      301 ttccgctatc ggctgaattt gattgcgagt gagatattta tgccagccag ccagacgcag
      361 acgcgccgag acagaactta atgggcccgc taacagcgcg atttgctggt gacccaatgc
      421 gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg gagaaaataa tactgttgat
      481 gggtgtctgg tcagagacat caagaaataa cgccggaaca ttagtgcagg cagcttccac
      541 agcaatggca tcctggtcat ccagcggata gttaatgatc agcccactga cgcgttgcgc
      601 gagaagattg tgcaccgccg ctttacaggc ttcgacgccg cttcgttcta ccatcgacac
      661 caccacgctg gcacccagtt gatcggcgcg agatttaatc gccgcgacaa tttgcgacgg
      721 cgcgtgcagg gccagactgg aggtggcaac gccaatcagc aacgactgtt tgcccgccag
      781 ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc atcgccgctt ccactttttc
      841 ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg cgggaaacgg tctgataaga
      901 gacaccggca tactctgcga catcgtataa cgttactggt ttcacattca ccaccctgaa
      961 ttgactctct tccgggcgct atcatgccat accgcgaaag gttttgcgcc attcgatggt
     1021 gtccgggatc tcgacgctct cccttatgcg actcctgcat taggaagcag cccagtagta
     1081 ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc atgcaaggag atggcgccca
     1141 acagtccccc ggccacgggg cctgccacca tacccacgcc gaaacaagcg ctcatgagcc
     1201 cgaagtggcg agcccgatct tccccatcgg tgatgtcggc gatataggcg ccagcaaccg
     1261 cacctgtggc gccggtgatg ccggccacga tgcgtccggc gtagaggatc gagatctcga
     1321 tcccgcgaaa ttaatacgac tcactatagg ggaattgtga gcggataaca attcccctct
     1381 agaaataatt ttgtttaact ttaagaagga gatataccat gggcagcagc catcatcatc
     1441 atcatcacga ttacgatatc ccaacgaccg aaaaccttta cttccagggc caatggcata
     1501 tgaaggaact taaatatgac gttctgataa tcggaggagg ttttgctggt tcttcagctg
     1561 cttaccaatt atcacgaaga ggtcttaaaa tattattagt agatagcaaa ccgtggaaca
     1621 ggataggaga caaaccttgt ggagatgcag taagtaaggc tcattttgat aagttaggta
     1681 tgccttatcc taaaggtgaa gagttagaaa ataaaataaa tggaataaag ctttacagcc
     1741 ctgatatgca gacagtatgg acagtaaatg gagagggttt tgaactaaat gccccactat
     1801 ataatcaaag agttcttaaa gaggcacagg atagaggagt cgaaatatgg gatttgacaa
     1861 cagcaatgaa acccatattt gaagatggtt atgtaaaagg agcagtcctc tttaatagaa
     1921 gaaccaatga ggaacttaca gtatactcta aggtagttgt tgaggctaca ggatattcaa
     1981 gaagtttcag aagtaaacta ccacctgaat tacccataac cgaagatcta gatgataagg
     2041 atgcagacgt agcgtacaga gaggttctac taactaagga agatatagag gaccatgact
     2101 acttgagaat attctttgat caagagacat ctccaggcgg ttattggtgg tacttcccta
     2161 aagggaagaa caaagtcaat gtcggtcttg gaattcaggg cggtatggga tacccaagca
     2221 ttcatgagta ttataaaaaa tatctagata agtatgcacc tgatgtggat aaaagtaaat
     2281 tattggttaa aggaggtgca ttggttccca ccagaagacc gttatataca atggcatgga
     2341 acggtataat agttatagga gattcagggt ttactgtaaa tcctgttcat ggcggaggaa
     2401 aaggttcagc catgatatct ggatattgtg cagcgaaagc tatactttca gcttttgaga
     2461 caggagattt ctcagcttca ggattatggg acatgaacat ttgctatgta aatgagtatg
     2521 gagcaaaaca agccagccta gacatattta ggaggttcct tcagaaactt agtaatgacg
     2581 acataaacta tggaatgaag aagaagatta taaaagaaga ggatctgctg gaagcaagtg
     2641 aaaaaggaga tcttcacttg tctgttgctg ataaggcaat gagagtaatc tctggtttag
     2701 gcagaccctc tttattattc aagctaaagg cggttgcaga gtctatgaag aaaataaagg
     2761 aattatatct gaattatcca agatccccct catctttagg tagttggaga agagaagttg
     2821 acaatgttct aacagagttt aacaaaagtt taagttagga tccgaattcg agctccgtcg
     2881 acaagcttgc ggccgcactc gagcaccacc accaccacca ctgagatccg gctgctaaca
     2941 aagcccgaaa ggaagctgag ttggctgctg ccaccgctga gcaataacta gcataacccc
     3001 ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa aggaggaact atatccggat
     3061 tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
     3121 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
     3181 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
     3241 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc
     3301 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
     3361 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc
     3421 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
     3481 acaaaaattt aacgcgaatt ttaacaaaat attaacgctt acaatttagg tggcactttt
     3541 cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat
     3601 ccgctcatga attaattctt agaaaaactc atcgagcatc aaatgaaact gcaatttatt
     3661 catatcagga ttatcaatac catatttttg aaaaagccgt ttctgtaatg aaggagaaaa
     3721 ctcaccgagg cagttccata ggatggcaag atcctggtat cggtctgcga ttccgactcg
     3781 tccaacatca atacaaccta ttaatttccc ctcgtcaaaa ataaggttat caagtgagaa
     3841 atcaccatga gtgacgactg aatccggtga gaatggcaaa agtttatgca tttctttcca
     3901 gacttgttca acaggccagc cattacgctc gtcatcaaaa tcactcgcat caaccaaacc
     3961 gttattcatt cgtgattgcg cctgagcgag acgaaatacg cgatcgctgt taaaaggaca
     4021 attacaaaca ggaatcgaat gcaaccggcg caggaacact gccagcgcat caacaatatt
     4081 ttcacctgaa tcaggatatt cttctaatac ctggaatgct gttttcccgg ggatcgcagt
     4141 ggtgagtaac catgcatcat caggagtacg gataaaatgc ttgatggtcg gaagaggcat
     4201 aaattccgtc agccagttta gtctgaccat ctcatctgta acatcattgg caacgctacc
     4261 tttgccatgt ttcagaaaca actctggcgc atcgggcttc ccatacaatc gatagattgt
     4321 cgcacctgat tgcccgacat tatcgcgagc ccatttatac ccatataaat cagcatccat
     4381 gttggaattt aatcgcggcc tagagcaaga cgtttcccgt tgaatatggc tcataacacc
     4441 ccttgtatta ctgtttatgt aagcagacag ttttattgtt catgaccaaa atcccttaac
     4501 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
     4561 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
     4621 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca
     4681 gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga
     4741 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca
     4801 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
     4861 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     4921 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     4981 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     5041 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     5101 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg
     5161 cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat
     5221 cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca
     5281 gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ctgatgcggt
     5341 attttctcct tacgcatctg tgcggtattt cacaccgcaa tggtgcactc tcagtacaat
     5401 ctgctctgat gccgcatagt taagccagta tacactccgc tatcgctacg tgactgggtc
     5461 atggctgcgc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
     5521 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
     5581 tcaccgtcat caccgaaacg cgcgaggcag ctgcggtaaa gctcatcagc gtggtcgtga
     5641 agcgattcac agatgtctgc ctgttcatcc gcgtccagct cgttgagttt ctccagaagc
     5701 gttaatgtct ggcttctgat aaagcgggcc atgttaaggg cggttttttc ctgtttggtc
     5761 actgatgcct ccgtgtaagg gggatttctg ttcatggggg taatgatacc gatgaaacga
     5821 gagaggatgc tcacgatacg ggttactgat gatgaacatg cccggttact ggaacgttgt
     5881 gagggtaaac aactggcggt atggatgcgg cgggaccaga gaaaaatcac tcagggtcaa
     5941 tgccagcgct tcgttaatac agatgtaggt gttccacagg gtagccagca gcatcctgcg
     6001 atgcagatcc ggaacataat ggtgcagggc gctgacttcc gcgtttccag actttacgaa
     6061 acacggaaac cgaagaccat tcatgttgtt gctcaggtcg cagacgtttt gcagcagcag
     6121 tcgcttcacg ttcgctcgcg tatcggtgat tcattctgct aaccagtaag gcaaccccgc
     6181 cagcctagcc gggtcctcaa cgacaggagc acgatcatgc gcacccgtgg ggccgccatg
     6241 ccggcgataa tggcctgctt ctcgccgaaa cgtttggtgg cgggaccagt gacgaaggct
     6301 tgagcgaggg cgtgcaagat tccgaatacc gcaagcgaca ggccgatcat cgtcgcgctc
     6361 cagcgaaagc ggtcctcgcc gaaaatgacc cagagcgctg ccggcacctg tcctacgagt
     6421 tgcatgataa agaagacagt cataagtgcg gcgacgatag tcatgccccg cgcccaccgg
     6481 aaggagctga ctgggttgaa ggctctcaag ggcatcggtc gagatcccgg tgcctaatga
     6541 gtgagctaac ttacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     6601 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     6661 cgccagggtg gtttttcttt tcaccagtga gacgggcaac agctgatt
//
