LOCUS       sequence-227284-        8673 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Luminescence-based glutaminase GAC isoform 3'UTR-mediated RNA 
            stability reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8673)
  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  JOURNAL   Unpublished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 8673)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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            "Unpublished"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 GAC
FEATURES             Location/Qualifiers
     source          1..8673
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..358
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        1..192
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(24..44)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      209..344
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     intron          428..560
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     543..562
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        605..623
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      627..636
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             633..1568
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     polyA_signal    4027..4075
                     /note="synthetic polyadenylation signal"
     promoter        4084..4835
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     regulatory      4865..4874
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4871..6523
                     /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             5830..5838
                     /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    6567..6688
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6578..6597)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6632..6651
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        6821..6925
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6926..6994,6995..7786)
                     /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(7144..7163)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7957..8545
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8446..8465
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
       61 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
      121 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
      181 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
      241 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc
      301 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
      361 gcttgattct tctgacacaa cagtctcgaa cttaagctgc agaagttggt cgtgaggcac
      421 tgggcaggta agtatcaagg ttacaagaca ggtttaagga gaccaataga aactgggctt
      481 gtcgagacag agaagactct tgcgtttctg ataggcacct attggtctta ctgacatcca
      541 ctttgccttt ctctccacag gtgtccactc ccagttcaat tacagctctt aaggctagag
      601 tacttaatac gactcactat aggctagcca ccatggcttc caaggtgtac gaccccgagc
      661 aacgcaaacg catgatcact gggcctcagt ggtgggctcg ctgcaagcaa atgaacgtgc
      721 tggactcctt catcaactac tatgattccg agaagcacgc cgagaacgcc gtgatttttc
      781 tgcatggtaa cgctgcctcc agctacctgt ggaggcacgt cgtgcctcac atcgagcccg
      841 tggctagatg catcatccct gatctgatcg gaatgggtaa gtccggcaag agcgggaatg
      901 gctcatatcg cctcctggat cactacaagt acctcaccgc ttggttcgag ctgctgaacc
      961 ttccaaagaa aatcatcttt gtgggccacg actggggggc ttgtctggcc tttcactact
     1021 cctacgagca ccaagacaag atcaaggcca tcgtccatgc tgagagtgtc gtggacgtga
     1081 tcgagtcctg ggacgagtgg cctgacatcg aggaggatat cgccctgatc aagagcgaag
     1141 agggcgagaa aatggtgctt gagaataact tcttcgtcga gaccatgctc ccaagcaaga
     1201 tcatgcggaa actggagcct gaggagttcg ctgcctacct ggagccattc aaggagaagg
     1261 gcgaggttag acggcctacc ctctcctggc ctcgcgagat ccctctcgtt aagggaggca
     1321 agcccgacgt cgtccagatt gtccgcaact acaacgccta ccttcgggcc agcgacgatc
     1381 tgcctaagat gttcatcgag tccgaccctg ggttcttttc caacgctatt gtcgagggag
     1441 ctaagaagtt ccctaacacc gagttcgtga aggtgaaggg cctccacttc agccaggagg
     1501 acgctccaga tgaaatgggt aagtacatca agagcttcgt ggagcgcgtg ctgaagaacg
     1561 agcagtaatt ctaggcgatc gctcgagaaa tgggttctag tttcagaatg tttcttcatt
     1621 taatctttca aacatcttta gctttttttt gcaagttata aatatttatt tgaggtattt
     1681 tttgttctca atcttgggtg ctggagccat aaagcttttt tttcctttta atctttgtat
     1741 aaaggcagta gattaagaag tgcatttgtt ggtctttaaa aagtatttac aagtacataa
     1801 atttgcttta tttttaaaaa tacaaaaagg aaaaatttaa attttttttg atgtaattaa
     1861 aatgttaact atgtggtcag ataatcccat tttacaatag taacagaaaa ttgtaattct
     1921 tagttctaaa attcacaaat taaactcata agttttgttg cattttgttt tttcttttcc
     1981 atttttaaaa ctaatgtgat gtctttagtg gcaatagaag gtacttctat gctaaataca
     2041 aaactaaaaa ggcaaaataa tgaaccccaa attattttat ttaaaatagc agtggattat
     2101 aaaattagct tgtgtttaca tttatgccat ttttggtgat agattggctt tacattttaa
     2161 aaaatttatt taaaaattta tcaaatgctt taaaatatga ctcctacttt ttttattttg
     2221 caactcctct gttctgtcag agttgttata tacaggagtg tcttatgtta ctaaaacatt
     2281 ccagccaaag aatttcagat gtgagataat gatgtttcat caataaaaag ctataatggt
     2341 tagttactca gaaggagaaa cagtgagtgt cttcaagtga attgttcacc taaacaattt
     2401 tattttcata ttatccacat aactttttct atgttatatt taaatatgaa tggcaaattt
     2461 tggtttttag cttttacatt ttattatctt aattttataa atgctaatat ttcttttgtg
     2521 ataagttata gcatctcata aagtttgttc tatttgaagt tttttagagt acttgagaaa
     2581 tgaatttagt ctgcaggtag taagtatgct actaaaatac gttagatcta aatcctttta
     2641 tttggtataa aaatgcaata ttgagaatca aaacttgttt ttaagagaac tatagattct
     2701 acacaacctg atttcaagta attattcata gtatttatag ttgtcttggc aaagtgattg
     2761 taaaattctg taggacctat tcacacttct tccttcttcc atatacttct ctggttttcc
     2821 ccatagttcc cctataattt caagtttgtt gaaacctgtt aattttagtg ggggattaga
     2881 agaaaaactt ggtggtttct tagcatgatg gtgtatgtat gtggtaatgg aaagtctgta
     2941 aaagtaaata tagtgtagca aaaaagattt cactgagtat tttagatact agtgcaaata
     3001 aagatagaaa atcttgatca taatgtctta agtttgggaa ctgtgatatt aagaaaagaa
     3061 attcccttct agaggtgctg gccaaaaagc cttttgggct aacttaagta ttaaatttat
     3121 atatttaaat aattatattt taagttgtag aggattttcc caaggatttt atgcttactt
     3181 gaatgttctt tgaatgttca gatgcatatc ctaactggat gcttctcaag gccttactgc
     3241 atatttgtgt tgcatattta tgttagttgc accagggcca tttgtagttt gggcaaccga
     3301 atgccttaat tggaaaaaag gcattgtggt ttcccctatg atctaaattg ttacatttta
     3361 ccatttcatt ccgaagttgg ttttacttta ttaaatgaag atttagtttt catatcgtat
     3421 acatagctgt atagatttca aaattaggtt gttaatttgt gtcacttact atttttgtgt
     3481 tggtaatgct ttaaatgcat acttaaaaat gaagtactgt tatctaagct actgtgttta
     3541 gaaaatgtta agaatgagca gaaattttta tagaaaagta taaacggaag aagagataag
     3601 atactgcgaa taggccctca aacttaaaaa agaaaaaact ttgccagttt taaggacata
     3661 ttttgattct ttcagtattc ttaacacctt tttaaacaaa gttcttgata gtacccacta
     3721 ttattgggtt tgttttatgc cattattgat tcttgatatt caagcattta caatgtagca
     3781 tatttgattt tcttttttct ttcttttttt ggcatcatta acatttcatt tgaaatgcat
     3841 attgttcttg aagtactttg tttttagcat aaatgttgtg cattttatct tagtgtttgg
     3901 atgaaaacat ttgtgttgtt tagctttcat ttgctttgta tatttaataa tgtaccttta
     3961 ttttccagta tgcctacatt ttgtattgca caataaattt attttaagct ggcggccgct
     4021 ggccgcaata aaatatcttt attttcatta catctgtgtg ttggtttttt gtgtgaggat
     4081 ctaaatgagt cttcggacct cgcgggggcc gcttaagcgg tggttagggt ttgtctgacg
     4141 cggggggagg gggaaggaac gaaacactct cattcggagg cggctcgggg tttggtcttg
     4201 gtggccacgg gcacgcagaa gagcgccgcg atcctcttaa gcaccccccc gccctccgtg
     4261 gaggcggggg tttggtcggc gggtggtaac tggcgggccg ctgactcggg cgggtcgcgc
     4321 gccccagagt gtgacctttt cggtctgctc gcagaccccc gggcggcgcc gccgcggcgg
     4381 cgacgggctc gctgggtcct aggctccatg gggaccgtat acgtggacag gctctggagc
     4441 atccgcacga ctgcggtgat attaccggag accttctgcg ggacgagccg ggtcacgcgg
     4501 ctgacgcgga gcgtccgttg ggcgacaaac accaggacgg ggcacaggta cactatcttg
     4561 tcacccggag gcgcgaggga ctgcaggagc ttcagggagt ggcgcagctg cttcatcccc
     4621 gtggcccgtt gctcgcgttt gctggcggtg tccccggaag aaatatattt gcatgtcttt
     4681 agttctatga tgacacaaac cccgcccagc gtcttgtcat tggcgaattc gaacacgcag
     4741 atgcagtcgg ggcggcgcgg tcccaggtcc acttcgcata ttaaggtgac gcgtgtggcc
     4801 tcgaacaccg agcgaccctg cagcgacccg cttaaaagct tggcattccg gtactgttgg
     4861 taaagccacc atggccgatg ctaagaacat taagaagggc cctgctccct tctaccctct
     4921 ggaggatggc accgctggcg agcagctgca caaggccatg aagaggtatg ccctggtgcc
     4981 tggcaccatt gccttcaccg atgcccacat tgaggtggac atcacctatg ccgagtactt
     5041 cgagatgtct gtgcgcctgg ccgaggccat gaagaggtac ggcctgaaca ccaaccaccg
     5101 catcgtggtg tgctctgaga actctctgca gttcttcatg ccagtgctgg gcgccctgtt
     5161 catcggagtg gccgtggccc ctgctaacga catttacaac gagcgcgagc tgctgaacag
     5221 catgggcatt tctcagccta ccgtggtgtt cgtgtctaag aagggcctgc agaagatcct
     5281 gaacgtgcag aagaagctgc ctatcatcca gaagatcatc atcatggact ctaagaccga
     5341 ctaccagggc ttccagagca tgtacacatt cgtgacatct catctgcctc ctggcttcaa
     5401 cgagtacgac ttcgtgccag agtctttcga cagggacaaa accattgccc tgatcatgaa
     5461 cagctctggg tctaccggcc tgcctaaggg cgtggccctg cctcatcgca ccgcctgtgt
     5521 gcgcttctct cacgcccgcg accctatttt cggcaaccag atcatccccg acaccgctat
     5581 tctgagcgtg gtgccattcc accacggctt cggcatgttc accaccctgg gctacctgat
     5641 ttgcggcttt cgggtggtgc tgatgtaccg cttcgaggag gagctgttcc tgcgcagcct
     5701 gcaagactac aaaattcagt ctgccctgct ggtgccaacc ctgttcagct tcttcgctaa
     5761 gagcaccctg atcgacaagt acgacctgtc taacctgcac gagattgcct ctggcggcgc
     5821 cccactgtct aaggaggtgg gcgaagccgt ggccaagcgc tttcatctgc caggcatccg
     5881 ccagggctac ggcctgaccg agacaaccag cgccattctg attaccccag agggcgacga
     5941 caagcctggc gccgtgggca aggtggtgcc attcttcgag gccaaggtgg tggacctgga
     6001 caccggcaag accctgggag tgaaccagcg cggcgagctg tgtgtgcgcg gccctatgat
     6061 tatgtccggc tacgtgaata accctgaggc cacaaacgcc ctgatcgaca aggacggctg
     6121 gctgcactct ggcgacattg cctactggga cgaggacgag cacttcttca tcgtggaccg
     6181 cctgaagtct ctgatcaagt acaagggcta ccaggtggcc ccagccgagc tggagtctat
     6241 cctgctgcag caccctaaca ttttcgacgc cggagtggcc ggcctgcccg acgacgatgc
     6301 cggcgagctg cctgccgccg tcgtcgtgct ggaacacggc aagaccatga ccgagaagga
     6361 gatcgtggac tatgtggcca gccaggtgac aaccgccaag aagctgcgcg gcggagtggt
     6421 gttcgtggac gaggtgccca agggcctgac cggcaagctg gacgcccgca agatccgcga
     6481 gatcctgatc aaggctaaga aaggcggcaa gatcgccgtg taataattct agagtcgggg
     6541 cggccggccg cttcgagcag acatgataag atacattgat gagtttggac aaaccacaac
     6601 tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt
     6661 aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca
     6721 ggttcagggg gaggtgtggg aggtttttta aagcaagtaa aacctctaca aatgtggtaa
     6781 aatcgataag gatccaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     6841 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     6901 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
     6961 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
     7021 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
     7081 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
     7141 tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg
     7201 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
     7261 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
     7321 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
     7381 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
     7441 aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt
     7501 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
     7561 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
     7621 atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa
     7681 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
     7741 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
     7801 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     7861 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     7921 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     7981 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     8041 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     8101 tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     8161 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     8221 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     8281 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     8341 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     8401 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     8461 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     8521 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     8581 cttttgctgg ccttttgctc acatggctcg acagatctgc gcagcaccat ggcctgaaat
     8641 aacctctgaa agaggaactt ggttaggtac ctt
//
