LOCUS       sequence-207683-        7521 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of glutaminase isoform KGA (wild type) with V5 
            and 6xHis tags.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7521)
  AUTHORS   Cassago A, Ferreira AP, Ferreira IM, Fornezari C, Gomes ER, Greene 
            KS, Pereira HM, Garratt RC, Dias SM, Ambrosio AL
  TITLE     Mitochondrial localization and structure-based phosphate activation 
            mechanism of Glutaminase C with implications for cancer metabolism.
  JOURNAL   Proc Natl Acad Sci U S A. 2012 Jan 24;109(4):1092-7. doi: 
            10.1073/pnas.1112495109. Epub 2012 Jan 6.
  PUBMED    22228304
REFERENCE   2  (bases 1 to 7521)
  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: 
            "10.1073/pnas.1112495109"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2012-01-24- 24"; volume: "109"; issue: "4"; pages: "1092-7"
COMMENT     Sequence Label: pcDNA3.1D V5-His-TOPO-KGA.wt
FEATURES             Location/Qualifiers
     source          1..7521
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             3018..3059
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             3069..3086
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3109..3126)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3115..3339
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3385..3813
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3472..3491)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3682..3703
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3822..3842)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3827..4157
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4008..4143
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4070..4089
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4224..5018
                     /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(4278..4297)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4888..4907
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5194..5315
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5231..5250)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5285..5304
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5364..5380)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5364..5380)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5377..5399)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5388..5404
                     /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(5412..5418,5419..5436,5437..5442))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5457..5478
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5595..5612)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5766..6354)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5846..5865)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6525..7316,7317..7385))
                     /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     7148..7167
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7386..7490)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagt
      901 taagcttggt accgagctcg gatccagtac ccttcaccat gatgcggctg cgaggctcgg
      961 ggatgctgcg ggacctgctc ctgcggtcgc ccgccggcgt gagcgcgact ctgcggcggg
     1021 cacagccctt ggtcaccctg tgccggcgtc cccgaggcgg gggacggccg gccgcgggcc
     1081 cggctgccgc cgcgcgactc cacccgtggt ggggcggggg cggctggccg gcggagcccc
     1141 tcgcgcgggg cctgtccagc tctccttcgg agatcttgca ggagctgggc aaggggagca
     1201 cgcatccgca gcccggggtg tcgccacccg ctgccccggc ggcgcccggc cccaaggacg
     1261 gccccgggga gacggacgcg tttggcaaca gcgagggcaa agagctggtg gcctcaggtg
     1321 aaaataaaat aaaacagggt ctgttaccta gcttggaaga tttgctgttc tatacaattg
     1381 ctgaaggaca agagaaaata cctgttcata aatttattac agcactcaaa tctacaggat
     1441 tgcgaacgtc tgatcccagg ttgaaagagt gtatggatat gttaagatta actcttcaaa
     1501 caacatcaga tggtgtcatg ctagacaaag atctttttaa aaaatgtgtt cagagcaaca
     1561 ttgttttgtt gacacaagca tttagaagaa agtttgtgat tcctgacttt atgtctttta
     1621 cctcacacat tgatgagtta tatgaaagtg ctaaaaagca gtctggagga aaggttgcag
     1681 attatattcc tcaactggcc aaattcagtc ccgatttgtg gggtgtgtct gtttgtacag
     1741 tagatggaca gaggcattct actggagata ccaaagttcc cttctgtctt cagtcctgtg
     1801 taaaaccttt gaaatatgcc attgctgtta atgatcttgg aactgaatat gtgcatcgat
     1861 atgttggaaa agagccgagt ggactaagat tcaacaaact atttttgaat gaagatgata
     1921 aaccacataa tcctatggta aatgctggag caattgttgt gacttcacta ataaagcaag
     1981 gagtaaataa tgctgaaaaa tttgactatg tcatgcagtt tttgaataag atggctggta
     2041 atgaatatgt tggattcagt aatgcaacgt ttcagtctga aagagaaagt ggagatcgaa
     2101 attttgcaat aggatattac ttaaaagaaa agaagtgttt tccagaaggc acagacatgg
     2161 ttggtatatt agacttctac ttccagctgt gctccattga agtgacttgt gaatcagcca
     2221 gtgtgatggc tgcgacactg gctaatggtg gtttctgccc aattactggt gaaagagtac
     2281 tgagccctga agcagttcga aatacattga gtttgatgca ttcctgtggc atgtatgact
     2341 tctcagggca gtttgctttc catgttggtc ttcctgcaaa atctggagtt gctgggggca
     2401 ttcttttagt tgtccccaat gttatgggta tgatgtgctg gtctcctcct ctggataaga
     2461 tgggcaacag tgttaaggga attcactttt gtcacgatct tgtttctctg tgtaatttcc
     2521 ataactatga taatttgaga cactttgcaa aaaaacttga tcctcgaaga gaaggtggtg
     2581 atcaaagggt aaagtcagtg ataaatcttt tgtttgctgc atatactgga gatgtgtctg
     2641 cacttcgaag atttgctttg tcagctatgg acatggaaca gcgggactat gattctagaa
     2701 cagcactcca tgtagctgct gcagagggtc atgttgaagt tgttaaattt ttgctggaag
     2761 cctgcaaagt aaaccctttc cccaaggaca ggtggaataa cactcccatg gatgaagcac
     2821 tgcactttgg acaccatgat gtatttaaaa ttctccaaga ataccaagtc cagtacacac
     2881 ctcaaggaga ttctgacaac gggaaggaaa atcaaaccgt ccataagaat cttgatggat
     2941 tgttgaaggg tcaagacaat tctgcagata tccagcacag tggcggccgc tcgagtctag
     3001 agggcccgcg gttcgaaggt aagcctatcc ctaaccctct cctcggtctc gattctacgc
     3061 gtaccggtca tcatcaccat caccattgag tttaaacccg ctgatcagcc tcgactgtgc
     3121 cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag
     3181 gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta
     3241 ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag
     3301 acaatagcag gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca
     3361 gctggggctc tagggggtat ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg
     3421 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg
     3481 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
     3541 gcatcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
     3601 agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
     3661 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta
     3721 tctcggtcta ttcttttgat ttataaggga ttttggggat ttcggcctat tggttaaaaa
     3781 atgagctgat ttaacaaaaa tttaacgcga attaattctg tggaatgtgt gtcagttagg
     3841 gtgtggaaag tccccaggct ccccaggcag gcagaagtat gcaaagcatg catctcaatt
     3901 agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     3961 tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa
     4021 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
     4081 aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag
     4141 gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcaag
     4201 agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg
     4261 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg
     4321 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc
     4381 tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga
     4441 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc
     4501 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag
     4561 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat
     4621 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg
     4681 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca
     4741 ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct
     4801 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg
     4861 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg
     4921 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc
     4981 gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgcgaa
     5041 atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc
     5101 tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc
     5161 ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt
     5221 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
     5281 agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct
     5341 agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     5401 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga
     5461 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     5521 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     5581 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     5641 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     5701 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     5761 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     5821 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     5881 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     5941 ggaagcgtgg cgctttctca atgctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     6001 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     6061 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     6121 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     6181 tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca
     6241 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     6301 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     6361 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     6421 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     6481 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     6541 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     6601 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     6661 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     6721 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     6781 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     6841 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     6901 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     6961 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     7021 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     7081 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     7141 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     7201 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     7261 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     7321 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     7381 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     7441 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     7501 cgaaaagtgc cacctgacgt c
//
