LOCUS       sequence-213697-        8803 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Luminescence-based glutaminase KGA isoform 3'UTR-mediated RNA 
            stability reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8803)
  AUTHORS   Sandra Martha Gomes Dias
  TITLE     HuR (ELAVL1) silencing sensitizes triple-negative breast cancer to 
            glutaminase-targeted therapy
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8803)
  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: 
            "Unpublished"
COMMENT     Sequence Label: psiCHECK2 3'UTR KGA
FEATURES             Location/Qualifiers
     source          1..8803
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      168..303
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     intron          387..519
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     502..521
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        564..582
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      586..595
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             592..1527
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     polyA_signal    4116..4164
                     /note="synthetic polyadenylation signal"
     promoter        4173..4924
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     regulatory      4954..4963
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4960..6612
                     /codon_start=1
                     /gene="luc+"
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="enhanced luc+ version of the luciferase gene"
                     /db_xref="GI:42718124"
                     /protein_id="AAS38485.1"
                     /translation="MADAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     RBS             5919..5927
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     polyA_signal    6656..6777
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6667..6686)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6721..6740
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        6910..7014
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7015..7083,7084..7875)
                     /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     complement(7233..7252)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8046..8634
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8535..8554
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        join(8763..8803,1..317)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        join(8763..8803,1..151)
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(join(8786..8803,1..3))
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
ORIGIN
        1 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
       61 agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
      121 tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa
      181 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
      241 aggccgaggc cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag
      301 gcctaggctt ttgcaaaaag cttgattctt ctgacacaac agtctcgaac ttaagctgca
      361 gaagttggtc gtgaggcact gggcaggtaa gtatcaaggt tacaagacag gtttaaggag
      421 accaatagaa actgggcttg tcgagacaga gaagactctt gcgtttctga taggcaccta
      481 ttggtcttac tgacatccac tttgcctttc tctccacagg tgtccactcc cagttcaatt
      541 acagctctta aggctagagt acttaatacg actcactata ggctagccac catggcttcc
      601 aaggtgtacg accccgagca acgcaaacgc atgatcactg ggcctcagtg gtgggctcgc
      661 tgcaagcaaa tgaacgtgct ggactccttc atcaactact atgattccga gaagcacgcc
      721 gagaacgccg tgatttttct gcatggtaac gctgcctcca gctacctgtg gaggcacgtc
      781 gtgcctcaca tcgagcccgt ggctagatgc atcatccctg atctgatcgg aatgggtaag
      841 tccggcaaga gcgggaatgg ctcatatcgc ctcctggatc actacaagta cctcaccgct
      901 tggttcgagc tgctgaacct tccaaagaaa atcatctttg tgggccacga ctggggggct
      961 tgtctggcct ttcactactc ctacgagcac caagacaaga tcaaggccat cgtccatgct
     1021 gagagtgtcg tggacgtgat cgagtcctgg gacgagtggc ctgacatcga ggaggatatc
     1081 gccctgatca agagcgaaga gggcgagaaa atggtgcttg agaataactt cttcgtcgag
     1141 accatgctcc caagcaagat catgcggaaa ctggagcctg aggagttcgc tgcctacctg
     1201 gagccattca aggagaaggg cgaggttaga cggcctaccc tctcctggcc tcgcgagatc
     1261 cctctcgtta agggaggcaa gcccgacgtc gtccagattg tccgcaacta caacgcctac
     1321 cttcgggcca gcgacgatct gcctaagatg ttcatcgagt ccgaccctgg gttcttttcc
     1381 aacgctattg tcgagggagc taagaagttc cctaacaccg agttcgtgaa ggtgaagggc
     1441 ctccacttca gccaggagga cgctccagat gaaatgggta agtacatcaa gagcttcgtg
     1501 gagcgcgtgc tgaagaacga gcagtaattc taggcgatcg ctcgaggaaa cggtctcaaa
     1561 tcccaagatt taaatcactt acctatttaa ttgtggaaaa tgattatgaa gaacatgtgt
     1621 atttctatct ggtagtgatg tatattttac atttgtcatt tcagtgttac tggagttttc
     1681 ttcattgtgc acacaggaca aatctgatct ctttgggaaa aaatagaaat aaaacaatct
     1741 ccctccataa tgtgagcaat attacctcgt gcattgtata atttgatgta aaagaaatag
     1801 ttaccaatgc tagcttgtgt ggtcttccat gatttatttg tgttttgtga attttcaatt
     1861 tatggtgatg atctgctgat atgcatttat aaagtaagct ctgttgtaca gtctgtccaa
     1921 atgggtcaag gttgccttta gaagcaaata gtgtgatttt caagacttca aatacaaatt
     1981 tagtttaagt gtttgaacaa ctatatgcac ttacggttgt gtgtttaaaa tgtctctctc
     2041 accccctagc ttcatgatgt gactcttaaa aaactataat agttaacaac tgttagtaag
     2101 atagaccaat tctgattaga ctttatcagg gaatctgttt aagatatgtt tggtgaccaa
     2161 aacgtatgtg tgaatgtagt tataatgctt ttgaaaaatt ttcctttttc tatatcccct
     2221 tagtccagcc tctcttctca gacatttagc tatctgcctc tttcctttag ctgggaaagt
     2281 gagagctggc atactatgca gtttttatgt tttccatagt aagtcagaaa atgcctccta
     2341 tttctggcat cagaactttg ccatttgtct acagaagacg aaccagagac aaaattacta
     2401 agtataaatt agtcaagttt atcagtctaa aaaacgaagg gatgtgcaac tgcagctctt
     2461 taagaagttt ttttttttta gcttctaggg taaagataaa ttcagaaatg ctctaagcta
     2521 ccaaagttat tctgaaagta tgggaactgc tacaactaac aaacatttgt ttccaagcct
     2581 gtcattaaga gtctgcatca agagatttgt cctccttggg ggaccactgg atcattccag
     2641 atttcttgtg atttttctat tgtgtaattc ttggtgggct ctgtagttta ataataagaa
     2701 aaaggccatt tcattttaaa ttgtgaccta taattctttg tcttgggttg gtaattcagg
     2761 attcatttgg aaagtgggta aaaggggctt caaaaaacgg atagaacagg attttctagg
     2821 agttacacat acattttatc ctgtcatacc tcgagataaa gtggcatgtt agtgaggagt
     2881 tctgatatta agcacacaca cacatgcaca caaatggact tctctgaagc tgtgtttagt
     2941 gaaatgagct caagtacatg aatgttagtt atcacataca gcaaattcct ttttttttct
     3001 ttttctatga gcacactctg ctgcttctaa actttacatg cctgatggca ccttactcca
     3061 gcagcctcca ggtgctttca ttttcacttc cagtctaagc cagtggctcc tgccactgcc
     3121 ctcccattac ctagatggca cctcctttgg tgaaaccacg gccaatgttc cttagctgca
     3181 ccaggcccga agctgttccc atgcttgagc ttccatgggg aggatgctga gtgagcagtt
     3241 tcctaccccg tggatctagc aagccatgga gacaggtagc atttgtaaga tgctgcacag
     3301 gagcagcatt atccccaaag atattacagg gtagacacgt tttaactgaa atcaatcaag
     3361 ataactttat tcaaagagca gcccgctttg tgtgactaaa atgaaacaag acagttgaat
     3421 tgtgtgactt gaagattacc aatgattttg aggcttttct ataataaaaa gaggttctaa
     3481 ccattatttg ggaacaaaga gagttttcat cttttttcag atcaaaacca ttctgtaaaa
     3541 tctttgttgt ttaattaaat gtgccgttat ttacccctga tgttatttat gactatgtgc
     3601 cgattcctgc tcgggctgtt tgctgttggc tggtaataat atatttgatt taaatgctgt
     3661 tgactgtgct attaactgct gccgtcagta aactccaaag atctttttgt tttggcttta
     3721 gtatcatatg tgctttttct gtatcctgag cgctctatat gatcatgtta atttaaagct
     3781 ttatacacat tgttgttttt gctggtctca tctttggtaa tatgctatac cccactgctg
     3841 cccgacactg ccctttagct gcagagctgg attagctgtt gaccatttga tgctgttgtc
     3901 tgtctggcag ggactgaatg acctgatgtc agatttagat tcttcctggg gattacacag
     3961 ctatgaatgt atttgcttct aaaacctccc aaagtgaatc taatcttaaa actacaagtt
     4021 gtaagtattc tgaaattggg aaacatttat tttaaatgca atcaggtagt gttgcttttt
     4081 acagcataat aaatatatgt gcggccgctg gccgcaataa aatatcttta ttttcattac
     4141 atctgtgtgt tggttttttg tgtgaggatc taaatgagtc ttcggacctc gcgggggccg
     4201 cttaagcggt ggttagggtt tgtctgacgc ggggggaggg ggaaggaacg aaacactctc
     4261 attcggaggc ggctcggggt ttggtcttgg tggccacggg cacgcagaag agcgccgcga
     4321 tcctcttaag cacccccccg ccctccgtgg aggcgggggt ttggtcggcg ggtggtaact
     4381 ggcgggccgc tgactcgggc gggtcgcgcg ccccagagtg tgaccttttc ggtctgctcg
     4441 cagacccccg ggcggcgccg ccgcggcggc gacgggctcg ctgggtccta ggctccatgg
     4501 ggaccgtata cgtggacagg ctctggagca tccgcacgac tgcggtgata ttaccggaga
     4561 ccttctgcgg gacgagccgg gtcacgcggc tgacgcggag cgtccgttgg gcgacaaaca
     4621 ccaggacggg gcacaggtac actatcttgt cacccggagg cgcgagggac tgcaggagct
     4681 tcagggagtg gcgcagctgc ttcatccccg tggcccgttg ctcgcgtttg ctggcggtgt
     4741 ccccggaaga aatatatttg catgtcttta gttctatgat gacacaaacc ccgcccagcg
     4801 tcttgtcatt ggcgaattcg aacacgcaga tgcagtcggg gcggcgcggt cccaggtcca
     4861 cttcgcatat taaggtgacg cgtgtggcct cgaacaccga gcgaccctgc agcgacccgc
     4921 ttaaaagctt ggcattccgg tactgttggt aaagccacca tggccgatgc taagaacatt
     4981 aagaagggcc ctgctccctt ctaccctctg gaggatggca ccgctggcga gcagctgcac
     5041 aaggccatga agaggtatgc cctggtgcct ggcaccattg ccttcaccga tgcccacatt
     5101 gaggtggaca tcacctatgc cgagtacttc gagatgtctg tgcgcctggc cgaggccatg
     5161 aagaggtacg gcctgaacac caaccaccgc atcgtggtgt gctctgagaa ctctctgcag
     5221 ttcttcatgc cagtgctggg cgccctgttc atcggagtgg ccgtggcccc tgctaacgac
     5281 atttacaacg agcgcgagct gctgaacagc atgggcattt ctcagcctac cgtggtgttc
     5341 gtgtctaaga agggcctgca gaagatcctg aacgtgcaga agaagctgcc tatcatccag
     5401 aagatcatca tcatggactc taagaccgac taccagggct tccagagcat gtacacattc
     5461 gtgacatctc atctgcctcc tggcttcaac gagtacgact tcgtgccaga gtctttcgac
     5521 agggacaaaa ccattgccct gatcatgaac agctctgggt ctaccggcct gcctaagggc
     5581 gtggccctgc ctcatcgcac cgcctgtgtg cgcttctctc acgcccgcga ccctattttc
     5641 ggcaaccaga tcatccccga caccgctatt ctgagcgtgg tgccattcca ccacggcttc
     5701 ggcatgttca ccaccctggg ctacctgatt tgcggctttc gggtggtgct gatgtaccgc
     5761 ttcgaggagg agctgttcct gcgcagcctg caagactaca aaattcagtc tgccctgctg
     5821 gtgccaaccc tgttcagctt cttcgctaag agcaccctga tcgacaagta cgacctgtct
     5881 aacctgcacg agattgcctc tggcggcgcc ccactgtcta aggaggtggg cgaagccgtg
     5941 gccaagcgct ttcatctgcc aggcatccgc cagggctacg gcctgaccga gacaaccagc
     6001 gccattctga ttaccccaga gggcgacgac aagcctggcg ccgtgggcaa ggtggtgcca
     6061 ttcttcgagg ccaaggtggt ggacctggac accggcaaga ccctgggagt gaaccagcgc
     6121 ggcgagctgt gtgtgcgcgg ccctatgatt atgtccggct acgtgaataa ccctgaggcc
     6181 acaaacgccc tgatcgacaa ggacggctgg ctgcactctg gcgacattgc ctactgggac
     6241 gaggacgagc acttcttcat cgtggaccgc ctgaagtctc tgatcaagta caagggctac
     6301 caggtggccc cagccgagct ggagtctatc ctgctgcagc accctaacat tttcgacgcc
     6361 ggagtggccg gcctgcccga cgacgatgcc ggcgagctgc ctgccgccgt cgtcgtgctg
     6421 gaacacggca agaccatgac cgagaaggag atcgtggact atgtggccag ccaggtgaca
     6481 accgccaaga agctgcgcgg cggagtggtg ttcgtggacg aggtgcccaa gggcctgacc
     6541 ggcaagctgg acgcccgcaa gatccgcgag atcctgatca aggctaagaa aggcggcaag
     6601 atcgccgtgt aataattcta gagtcggggc ggccggccgc ttcgagcaga catgataaga
     6661 tacattgatg agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt
     6721 gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa acaagttaac
     6781 aacaacaatt gcattcattt tatgtttcag gttcaggggg aggtgtggga ggttttttaa
     6841 agcaagtaaa acctctacaa atgtggtaaa atcgataagg atccaggtgg cacttttcgg
     6901 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     6961 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt
     7021 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
     7081 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
     7141 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
     7201 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt
     7261 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     7321 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     7381 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     7441 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
     7501 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta
     7561 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     7621 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     7681 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
     7741 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     7801 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     7861 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
     7921 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     7981 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     8041 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     8101 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     8161 ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac
     8221 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     8281 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     8341 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     8401 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     8461 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     8521 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     8581 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     8641 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catggctcga
     8701 cagatctgcg cagcaccatg gcctgaaata acctctgaaa gaggaacttg gttaggtacc
     8761 ttctgaggcg gaaagaacca gctgtggaat gtgtgtcagt tag
//
