LOCUS       sequence-258707-        6763 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses R132 mutant IDH1 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6763)
  AUTHORS   Zhao S, Lin Y, Xu W, Jiang W, Zha Z, Wang P, Yu W, Li Z, Gong L, 
            Peng Y, Ding J, Lei Q, Guan KL, Xiong Y
  TITLE     Glioma-derived mutations in IDH1 dominantly inhibit IDH1 catalytic 
            activity and induce HIF-1alpha.
  JOURNAL   Science. 2009 Apr 10;324(5924):261-5. doi: 10.1126/science.1170944.
  PUBMED    19359588
REFERENCE   2  (bases 1 to 6763)
  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"; doi: "10.1126/science"; 
            journalName: "Science"; date: "2009-04-10- 10"; volume: "324"; 
            issue: "5924"; pages: "261-5"
COMMENT     Sequence Label: pcDNA3-Flag-IDH1-R132H
FEATURES             Location/Qualifiers
     source          1..6763
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1977..2000
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2014..2043
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2059..2076
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2120..2137)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2126..2350
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2396..2824
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2483..2502)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2693..2714
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2833..2853)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2838..3167
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3018..3153
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3080..3099
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3234..4028
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3288..3307)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3898..3917
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4202..4323
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4239..4258)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4293..4312
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4372..4388)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4372..4388)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4385..4407)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4396..4412
                     /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)."
     promoter        complement(join(4420..4426,4427..4444,4445..4450))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4465..4486
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4603..4620)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4774..5362)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4854..4873)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5533..6324,6325..6393))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6156..6175
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6394..6498)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6573..6592)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctg
      661 gctagcgttt aaacgggccc tctagactcg agcggccgcc actgtgctgg atatctgcag
      721 aattcatgtc caaaaaaatc agtggcggtt ctgtggtaga gatgcaagga gatgaaatga
      781 cacgaatcat ttgggaattg attaaagaga aactcatttt tccctacgtg gaattggatc
      841 tacatagcta tgatttaggc atagagaatc gtgatgccac caacgaccaa gtcaccaagg
      901 atgccgcaga agctataaag aagcataatg ttggcgtcaa atgtgccact atcactcctg
      961 atgagaagag ggttgaggag ttcaagttga aacaaatgtg gaaatcacca aatggcacca
     1021 tacgaaatat tctgggtggc acggtcttca gagaagccat tatctgcaaa aatatccccc
     1081 ggcttgtgag tggatgggta aaacctatca tcataggtca tcatgcttat ggggatcaat
     1141 acagagcaac tgattttgtt gttcctgggc ctggaaaagt agagataacc tacacaccaa
     1201 gtgacggaac ccaaaaggtg acatacctgg tacataactt tgaagaaggt ggtggtgttg
     1261 ccatggggat gtataatcaa gataagtcaa ttgaagattt tgcacacagt tccttccaaa
     1321 tggctctgtc taagggttgg cctttgtatc tgagcaccaa aaacactatt ctgaagaaat
     1381 atgatgggcg ttttaaagac atctttcagg agatatatga caagcagtac aagtcccagt
     1441 ttgaagctca aaagatctgg tatgagcata ggctcatcga cgacatggtg gcccaagcta
     1501 tgaaatcaga gggaggcttc atctgggcct gtaaaaacta tgatggtgac gtgcagtcgg
     1561 actctgtggc ccaagggtat ggctctctcg gcatgatgac cagcgtgctg gtttgtccag
     1621 atggcaagac agtagaagca gaggctgccc acgggactgt aacccgacac taccgcatgt
     1681 accagaaagg acaggagacg tccaccaatc ccattgcttc catttttgcc tggaccagag
     1741 ggttagccca cagagcaaag cttgataaca ataaagagct tgccttcttt gcaaatgctt
     1801 tggaagaagt ctctattgag acaattgagg ctggcttcat gaccaaggac ttggctgctt
     1861 gcattaaagg tttacccaat gtgcaacgtt ctgactactt gaatacattt gagttcatgg
     1921 ataaacttgg agaaaacttg aagatcaaac tagctcaggc caaacttgtg gatcccgatt
     1981 acaaggatga cgacgataag tagaagcttt ctagaacaaa aactcatctc agaagaggat
     2041 ctgaatagcg ccgtcgacca tcatcatcat catcattgag tttaaacggt ctccagctta
     2101 agtttaaacc gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc
     2161 ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa
     2221 aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg
     2281 gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg
     2341 ggctctatgg cttctgaggc ggaaagaacc agctggggct ctagggggta tccccacgcg
     2401 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     2461 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     2521 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     2581 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg
     2641 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     2701 ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg
     2761 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     2821 aattaattct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag
     2881 gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag
     2941 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc
     3001 cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc
     3061 atggctgact aatttttttt atttatgcag aggccgaggc cgcctctgcc tctgagctat
     3121 tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctcccgggag
     3181 cttgtatatc cattttcgga tctgatcaag agacaggatg aggatcgttt cgcatgattg
     3241 aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg
     3301 actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg
     3361 ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg
     3421 aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg
     3481 ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc
     3541 tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc
     3601 tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc
     3661 gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc
     3721 aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg
     3781 atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct
     3841 tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt
     3901 tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc
     3961 tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt
     4021 tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca acctgccatc
     4081 acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg
     4141 ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc
     4201 caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     4261 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     4321 ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct
     4381 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     4441 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     4501 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     4561 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     4621 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     4681 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     4741 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     4801 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     4861 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     4921 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     4981 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     5041 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     5101 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     5161 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     5221 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     5281 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     5341 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     5401 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     5461 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     5521 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     5581 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     5641 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     5701 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     5761 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     5821 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     5881 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     5941 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     6001 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     6061 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     6121 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     6181 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     6241 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     6301 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     6361 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     6421 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     6481 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg acggatcggg
     6541 agatctcccg atcccctatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     6601 agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt
     6661 taagctacaa caaggcaagg cttgaccgac aattgcatga agaatctgct tagggttagg
     6721 cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc gtt
//
