LOCUS       sequence-207792-        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  JOURNAL   Unpublished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            "Unpublished"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"
     promoter        62..419
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        62..253
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(85..105)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      270..405
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     intron          489..621
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     604..623
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        666..684
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      688..697
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             694..1629
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     polyA_signal    4218..4266
                     /note="synthetic polyadenylation signal"
     promoter        4275..5026
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     regulatory      5056..5065
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             5062..6714
                     /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             6021..6029
                     /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    6758..6879
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6769..6788)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6823..6842
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        7012..7116
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7117..7185,7186..7977)
                     /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(7335..7354)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8148..8736
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8637..8656
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 agatctgcgc agcaccatgg cctgaaataa cctctgaaag aggaacttgg ttaggtacct
       61 tctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
      121 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
      181 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
      241 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
      301 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg
      361 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa
      421 agcttgattc ttctgacaca acagtctcga acttaagctg cagaagttgg tcgtgaggca
      481 ctgggcaggt aagtatcaag gttacaagac aggtttaagg agaccaatag aaactgggct
      541 tgtcgagaca gagaagactc ttgcgtttct gataggcacc tattggtctt actgacatcc
      601 actttgcctt tctctccaca ggtgtccact cccagttcaa ttacagctct taaggctaga
      661 gtacttaata cgactcacta taggctagcc accatggctt ccaaggtgta cgaccccgag
      721 caacgcaaac gcatgatcac tgggcctcag tggtgggctc gctgcaagca aatgaacgtg
      781 ctggactcct tcatcaacta ctatgattcc gagaagcacg ccgagaacgc cgtgattttt
      841 ctgcatggta acgctgcctc cagctacctg tggaggcacg tcgtgcctca catcgagccc
      901 gtggctagat gcatcatccc tgatctgatc ggaatgggta agtccggcaa gagcgggaat
      961 ggctcatatc gcctcctgga tcactacaag tacctcaccg cttggttcga gctgctgaac
     1021 cttccaaaga aaatcatctt tgtgggccac gactgggggg cttgtctggc ctttcactac
     1081 tcctacgagc accaagacaa gatcaaggcc atcgtccatg ctgagagtgt cgtggacgtg
     1141 atcgagtcct gggacgagtg gcctgacatc gaggaggata tcgccctgat caagagcgaa
     1201 gagggcgaga aaatggtgct tgagaataac ttcttcgtcg agaccatgct cccaagcaag
     1261 atcatgcgga aactggagcc tgaggagttc gctgcctacc tggagccatt caaggagaag
     1321 ggcgaggtta gacggcctac cctctcctgg cctcgcgaga tccctctcgt taagggaggc
     1381 aagcccgacg tcgtccagat tgtccgcaac tacaacgcct accttcgggc cagcgacgat
     1441 ctgcctaaga tgttcatcga gtccgaccct gggttctttt ccaacgctat tgtcgaggga
     1501 gctaagaagt tccctaacac cgagttcgtg aaggtgaagg gcctccactt cagccaggag
     1561 gacgctccag atgaaatggg taagtacatc aagagcttcg tggagcgcgt gctgaagaac
     1621 gagcagtaat tctaggcgat cgctcgagga aacggtctca aatcccaaga tttaaatcac
     1681 ttacctattt aattgtggaa aatgattatg aagaacatgt gtatttctat ctggtagtga
     1741 tgtatatttt acatttgtca tttcagtgtt actggagttt tcttcattgt gcacacagga
     1801 caaatctgat ctctttggga aaaaatagaa ataaaacaat ctccctccat aatgtgagca
     1861 atattacctc gtgcattgta taatttgatg taaaagaaat agttaccaat gctagcttgt
     1921 gtggtcttcc atgatttatt tgtgttttgt gaattttcaa tttatggtga tgatctgctg
     1981 atatgcattt ataaagtaag ctctgttgta cagtctgtcc aaatgggtca aggttgcctt
     2041 tagaagcaaa tagtgtgatt ttcaagactt caaatacaaa tttagtttaa gtgtttgaac
     2101 aactatatgc acttacggtt gtgtgtttaa aatgtctctc tcacccccta gcttcatgat
     2161 gtgactctta aaaaactata atagttaaca actgttagta agatagacca attctgatta
     2221 gactttatca gggaatctgt ttaagatatg tttggtgacc aaaacgtatg tgtgaatgta
     2281 gttataatgc ttttgaaaaa ttttcctttt tctatatccc cttagtccag cctctcttct
     2341 cagacattta gctatctgcc tctttccttt agctgggaaa gtgagagctg gcatactatg
     2401 cagtttttat gttttccata gtaagtcaga aaatgcctcc tatttctggc atcagaactt
     2461 tgccatttgt ctacagaaga cgaaccagag acaaaattac taagtataaa ttagtcaagt
     2521 ttatcagtct aaaaaacgaa gggatgtgca actgcagctc tttaagaagt tttttttttt
     2581 tagcttctag ggtaaagata aattcagaaa tgctctaagc taccaaagtt attctgaaag
     2641 tatgggaact gctacaacta acaaacattt gtttccaagc ctgtcattaa gagtctgcat
     2701 caagagattt gtcctccttg ggggaccact ggatcattcc agatttcttg tgatttttct
     2761 attgtgtaat tcttggtggg ctctgtagtt taataataag aaaaaggcca tttcatttta
     2821 aattgtgacc tataattctt tgtcttgggt tggtaattca ggattcattt ggaaagtggg
     2881 taaaaggggc ttcaaaaaac ggatagaaca ggattttcta ggagttacac atacatttta
     2941 tcctgtcata cctcgagata aagtggcatg ttagtgagga gttctgatat taagcacaca
     3001 cacacatgca cacaaatgga cttctctgaa gctgtgttta gtgaaatgag ctcaagtaca
     3061 tgaatgttag ttatcacata cagcaaattc cttttttttt ctttttctat gagcacactc
     3121 tgctgcttct aaactttaca tgcctgatgg caccttactc cagcagcctc caggtgcttt
     3181 cattttcact tccagtctaa gccagtggct cctgccactg ccctcccatt acctagatgg
     3241 cacctccttt ggtgaaacca cggccaatgt tccttagctg caccaggccc gaagctgttc
     3301 ccatgcttga gcttccatgg ggaggatgct gagtgagcag tttcctaccc cgtggatcta
     3361 gcaagccatg gagacaggta gcatttgtaa gatgctgcac aggagcagca ttatccccaa
     3421 agatattaca gggtagacac gttttaactg aaatcaatca agataacttt attcaaagag
     3481 cagcccgctt tgtgtgacta aaatgaaaca agacagttga attgtgtgac ttgaagatta
     3541 ccaatgattt tgaggctttt ctataataaa aagaggttct aaccattatt tgggaacaaa
     3601 gagagttttc atcttttttc agatcaaaac cattctgtaa aatctttgtt gtttaattaa
     3661 atgtgccgtt atttacccct gatgttattt atgactatgt gccgattcct gctcgggctg
     3721 tttgctgttg gctggtaata atatatttga tttaaatgct gttgactgtg ctattaactg
     3781 ctgccgtcag taaactccaa agatcttttt gttttggctt tagtatcata tgtgcttttt
     3841 ctgtatcctg agcgctctat atgatcatgt taatttaaag ctttatacac attgttgttt
     3901 ttgctggtct catctttggt aatatgctat accccactgc tgcccgacac tgccctttag
     3961 ctgcagagct ggattagctg ttgaccattt gatgctgttg tctgtctggc agggactgaa
     4021 tgacctgatg tcagatttag attcttcctg gggattacac agctatgaat gtatttgctt
     4081 ctaaaacctc ccaaagtgaa tctaatctta aaactacaag ttgtaagtat tctgaaattg
     4141 ggaaacattt attttaaatg caatcaggta gtgttgcttt ttacagcata ataaatatat
     4201 gtgcggccgc tggccgcaat aaaatatctt tattttcatt acatctgtgt gttggttttt
     4261 tgtgtgagga tctaaatgag tcttcggacc tcgcgggggc cgcttaagcg gtggttaggg
     4321 tttgtctgac gcggggggag ggggaaggaa cgaaacactc tcattcggag gcggctcggg
     4381 gtttggtctt ggtggccacg ggcacgcaga agagcgccgc gatcctctta agcacccccc
     4441 cgccctccgt ggaggcgggg gtttggtcgg cgggtggtaa ctggcgggcc gctgactcgg
     4501 gcgggtcgcg cgccccagag tgtgaccttt tcggtctgct cgcagacccc cgggcggcgc
     4561 cgccgcggcg gcgacgggct cgctgggtcc taggctccat ggggaccgta tacgtggaca
     4621 ggctctggag catccgcacg actgcggtga tattaccgga gaccttctgc gggacgagcc
     4681 gggtcacgcg gctgacgcgg agcgtccgtt gggcgacaaa caccaggacg gggcacaggt
     4741 acactatctt gtcacccgga ggcgcgaggg actgcaggag cttcagggag tggcgcagct
     4801 gcttcatccc cgtggcccgt tgctcgcgtt tgctggcggt gtccccggaa gaaatatatt
     4861 tgcatgtctt tagttctatg atgacacaaa ccccgcccag cgtcttgtca ttggcgaatt
     4921 cgaacacgca gatgcagtcg gggcggcgcg gtcccaggtc cacttcgcat attaaggtga
     4981 cgcgtgtggc ctcgaacacc gagcgaccct gcagcgaccc gcttaaaagc ttggcattcc
     5041 ggtactgttg gtaaagccac catggccgat gctaagaaca ttaagaaggg ccctgctccc
     5101 ttctaccctc tggaggatgg caccgctggc gagcagctgc acaaggccat gaagaggtat
     5161 gccctggtgc ctggcaccat tgccttcacc gatgcccaca ttgaggtgga catcacctat
     5221 gccgagtact tcgagatgtc tgtgcgcctg gccgaggcca tgaagaggta cggcctgaac
     5281 accaaccacc gcatcgtggt gtgctctgag aactctctgc agttcttcat gccagtgctg
     5341 ggcgccctgt tcatcggagt ggccgtggcc cctgctaacg acatttacaa cgagcgcgag
     5401 ctgctgaaca gcatgggcat ttctcagcct accgtggtgt tcgtgtctaa gaagggcctg
     5461 cagaagatcc tgaacgtgca gaagaagctg cctatcatcc agaagatcat catcatggac
     5521 tctaagaccg actaccaggg cttccagagc atgtacacat tcgtgacatc tcatctgcct
     5581 cctggcttca acgagtacga cttcgtgcca gagtctttcg acagggacaa aaccattgcc
     5641 ctgatcatga acagctctgg gtctaccggc ctgcctaagg gcgtggccct gcctcatcgc
     5701 accgcctgtg tgcgcttctc tcacgcccgc gaccctattt tcggcaacca gatcatcccc
     5761 gacaccgcta ttctgagcgt ggtgccattc caccacggct tcggcatgtt caccaccctg
     5821 ggctacctga tttgcggctt tcgggtggtg ctgatgtacc gcttcgagga ggagctgttc
     5881 ctgcgcagcc tgcaagacta caaaattcag tctgccctgc tggtgccaac cctgttcagc
     5941 ttcttcgcta agagcaccct gatcgacaag tacgacctgt ctaacctgca cgagattgcc
     6001 tctggcggcg ccccactgtc taaggaggtg ggcgaagccg tggccaagcg ctttcatctg
     6061 ccaggcatcc gccagggcta cggcctgacc gagacaacca gcgccattct gattacccca
     6121 gagggcgacg acaagcctgg cgccgtgggc aaggtggtgc cattcttcga ggccaaggtg
     6181 gtggacctgg acaccggcaa gaccctggga gtgaaccagc gcggcgagct gtgtgtgcgc
     6241 ggccctatga ttatgtccgg ctacgtgaat aaccctgagg ccacaaacgc cctgatcgac
     6301 aaggacggct ggctgcactc tggcgacatt gcctactggg acgaggacga gcacttcttc
     6361 atcgtggacc gcctgaagtc tctgatcaag tacaagggct accaggtggc cccagccgag
     6421 ctggagtcta tcctgctgca gcaccctaac attttcgacg ccggagtggc cggcctgccc
     6481 gacgacgatg ccggcgagct gcctgccgcc gtcgtcgtgc tggaacacgg caagaccatg
     6541 accgagaagg agatcgtgga ctatgtggcc agccaggtga caaccgccaa gaagctgcgc
     6601 ggcggagtgg tgttcgtgga cgaggtgccc aagggcctga ccggcaagct ggacgcccgc
     6661 aagatccgcg agatcctgat caaggctaag aaaggcggca agatcgccgt gtaataattc
     6721 tagagtcggg gcggccggcc gcttcgagca gacatgataa gatacattga tgagtttgga
     6781 caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt
     6841 gctttatttg taaccattat aagctgcaat aaacaagtta acaacaacaa ttgcattcat
     6901 tttatgtttc aggttcaggg ggaggtgtgg gaggtttttt aaagcaagta aaacctctac
     6961 aaatgtggta aaatcgataa ggatccaggt ggcacttttc ggggaaatgt gcgcggaacc
     7021 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     7081 tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc
     7141 gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg
     7201 gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat
     7261 ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc
     7321 acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa
     7381 ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa
     7441 aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt
     7501 gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct
     7561 tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat
     7621 gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg
     7681 cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg
     7741 atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt
     7801 attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg
     7861 ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg
     7921 gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg
     7981 tcagaccaag tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa
     8041 aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt
     8101 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt
     8161 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
     8221 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag
     8281 ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta
     8341 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
     8401 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
     8461 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
     8521 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
     8581 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
     8641 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
     8701 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
     8761 cggttcctgg ccttttgctg gccttttgct cacatggctc gac
//
