LOCUS       sequence-176863-        6776 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expresses IDH2 (R172K mutant) in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6776)
  AUTHORS   Showalter MR, Hatakeyama J, Cajka T, VanderVorst K, Carraway KL, 
            Fiehn O
  TITLE     Replication Study: The common feature of leukemia-associated IDH1 
            and IDH2 mutations is a neomorphic enzyme activity converting 
            alpha-ketoglutarate to 2-hydroxyglutarate.
  JOURNAL   Elife. 2017 Jun 27;6. pii: e26030. doi: 10.7554/eLife.26030.
  PUBMED    28653623
REFERENCE   2  (bases 1 to 6776)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Elife."; 
            date: "2017-06-27"; volume: "6. pii: e26030. doi"; pages: " 
            10.7554/eLife.26030"
COMMENT     Sequence Label: pcDNA3.1-IDH2-R172K
FEATURES             Location/Qualifiers
     source          1..6776
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(16..36)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        21..350
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      201..336
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     263..282
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             417..1211
                     /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(471..490)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1081..1100
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    1385..1506
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1422..1441)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1476..1495
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1555..1571)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1555..1571)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1568..1590)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1579..1595
                     /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(1603..1609,1610..1627,1628..1633))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1648..1669
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1786..1803)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1957..2545)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2037..2056)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2716..3507,3508..3576))
                     /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     3339..3358
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3577..3681)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3756..3775)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        3947..4326
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4327..4530
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4481..4501
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4575..4594
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4575..4593
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6079..6096)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6085..6309
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      join(6355..6776,1..7)
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6442..6461)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6652..6673
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag
       61 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc
      121 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag
      181 tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc
      241 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctct gcctctgagc
      301 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcccgg
      361 gagcttgtat atccattttc ggatctgatc aagagacagg atgaggatcg tttcgcatga
      421 ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct
      481 atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc
      541 aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcagg
      601 acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg
      661 acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc
      721 tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc
      781 ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg
      841 agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc
      901 atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg
      961 aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc
     1021 gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag
     1081 cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg
     1141 tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg
     1201 agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc
     1261 atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt
     1321 ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca
     1381 ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     1441 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     1501 atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata
     1561 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     1621 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     1681 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     1741 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     1801 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     1861 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     1921 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     1981 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     2041 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     2101 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     2161 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     2221 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     2281 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     2341 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     2401 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     2461 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     2521 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     2581 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     2641 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     2701 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     2761 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     2821 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     2881 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     2941 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     3001 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     3061 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     3121 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     3181 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     3241 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     3301 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     3361 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     3421 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     3481 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     3541 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     3601 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     3661 taaacaaata ggggtttcgc gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc
     3721 gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga tgccgcatag
     3781 ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa
     3841 atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct gcttagggtt
     3901 aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgttgaca ttgattattg
     3961 actagttatt aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc
     4021 cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca
     4081 ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt
     4141 caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg
     4201 ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag
     4261 tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt
     4321 accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg
     4381 ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa
     4441 cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt
     4501 gtacggtggg aggtctatat aagcagagct ctctggctaa ctagagaacc cactgcttac
     4561 tggcttatcg aaattaatac gactcactat agggagaccc aagctggcta gcgtttaaac
     4621 ttaagcttgg taccgagctc ggatccacca tggccggcta cctgcgggtc gtgcgctcgc
     4681 tctgcagagc ctcaggctcg cggccggcct gggcgccggc ggccctgaca gcccccacct
     4741 cgcaagagca gccgcggcgc cactatgccg acaaaaggat caaggtggcg aagcccgtgg
     4801 tggagatgga tggtgatgag atgacccgta ttatctggca gttcatcaag gagaagctca
     4861 tcctgcccca cgtggacatc cagctaaagt attttgacct cgggctccca aaccgtgacc
     4921 agactgatga ccaggtcacc attgactctg cactggccac ccagaagtac agtgtggctg
     4981 tcaagtgtgc caccatcacc cctgatgagg cccgtgtgga agagttcaag ctgaagaaga
     5041 tgtggaaaag tcccaatgga actatccgga acatcctggg ggggactgtc ttccgggagc
     5101 ccatcatctg caaaaacatc ccacgcctag tccctggctg gaccaagccc atcaccattg
     5161 gcaagcacgc ccatggcgac cagtacaagg ccacagactt tgtggcagac cgggccggca
     5221 ctttcaaaat ggtcttcacc ccaaaagatg gcagtggtgt caaggagtgg gaagtgtaca
     5281 acttccccgc aggcggcgtg ggcatgggca tgtacaacac cgacgagtcc atctcaggtt
     5341 ttgcgcacag ctgcttccag tatgccatcc agaagaaatg gccgctgtac atgagcacca
     5401 agaacaccat actgaaagcc tacgatgggc gtttcaagga catcttccag gagatctttg
     5461 acaagcacta taagaccgac ttcgacaaga ataagatctg gtatgagcac cggctcattg
     5521 atgacatggt ggctcaggtc ctcaagtctt cgggtggctt tgtgtgggcc tgcaagaact
     5581 atgacggaga tgtgcagtca gacatcctgg cccagggctt tggctccctt ggcctgatga
     5641 cgtccgtcct ggtctgccct gatgggaaga cgattgaggc tgaggccgct catgggaccg
     5701 tcacccgcca ctatcgggag caccagaagg gccggcccac cagcaccaac cccatcgcca
     5761 gcatctttgc ctggacacgt ggcctggagc accgggggaa gctggatggg aaccaagacc
     5821 tcatcaggtt tgcccagatg ctggagaagg tgtgcgtgga gacggtggag agtggagcca
     5881 tgaccaagga cctggcgggc tgcattcacg gcctcagcaa tgtgaagctg aacgagcact
     5941 tcctgaacac cacggacttc ctcgacacca tcaagagcaa cctggacaga gccctgggca
     6001 ggcagtaaga attctgcaga tatccagcac agtggcggcc gctcgagtct agagggcccg
     6061 tttaaacccg ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc
     6121 cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa
     6181 atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg
     6241 ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg
     6301 gctctatggc ttctgaggcg gaaagaacca gctggggctc tagggggtat ccccacgcgc
     6361 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     6421 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     6481 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     6541 tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc
     6601 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     6661 tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga
     6721 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaac
//
