LOCUS       sequence-213705-        8700 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  gamma-Retroviral transfer vector for expressing HuR (wild type), V5 
            and mKO2 tags, IRES-driven Puromycin selection..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8700)
  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  JOURNAL   Unpublished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 8700)
  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            "Unpublished"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: pQC V5 mKO2 HuR.wt IRES Puro
FEATURES             Location/Qualifiers
     source          1..8700
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             585..760
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     misc_feature    823..1022
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1223..1639
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1569..1591
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1607..1623
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     enhancer        1676..1979
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1980..2183
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2180..2204
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             2338..2379
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2389..3042
                     /codon_start=1
                     /product="monomeric Kusabira-Orange 2 fluorescent protein"
                     /label=mKO2
                     /translation="MVSVIKPEMKMRYYMDGSVNGHEFTIEGEGTGRPYEGHQEMTLRV
                     TMAEGGPMPFAFDLVSHVFCYGHRVFTKYPEEIPDYFKQAFPEGLSWERSLEFEDGGSA
                     SVSAHISLRGNTFYHKSKFTGVNFPADGPIMQNQSVDWEPSTEKITASDGVLKGDVTMY
                     LKLEGGGNHKCQMKTTYKAAKEILEMPGDHYIGHRLVRKTEGNITEQVEDAVAHS"
     CDS             2389..3042
                     /codon_start=1
                     /product="humanized monomeric Kusabira-Orange fluorescent 
                     protein"
                     /label=hmKO
                     /note="mammalian codon-optimized"
                     /translation="MVSVIKPEMKMRYYMDGSVNGHEFTIEGEGTGRPYEGHQEMTLRV
                     TMAEGGPMPFAFDLVSHVFCYGHRVFTKYPEEIPDYFKQAFPEGLSWERSLEFEDGGSA
                     SVSAHISLRGNTFYHKSKFTGVNFPADGPIMQNQSVDWEPSTEKITASDGVLKGDVTMY
                     LKLEGGGNHKCQMKTTYKAAKEILEMPGDHYIGHRLVRKTEGNITEQVEDAVAHS"
     misc_feature    4096..4669
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4263..4280)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     4490..4509
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4710..5309
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4710..4729)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5206..5226
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             5478..5903
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     Moloney murine leukemia virus"
     primer_bind     complement(6006..6028)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6164..6184)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6169..6498
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6349..6484
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6520..6539
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(6620..6637)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6791..7379)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6871..6890)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7550..8341,8342..8410))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     8173..8192
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8411..8515)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     8583..8601
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcaataa aagagcccac
      601 aacccctcac tcggcgcgcc agtcttccga tagactgcgt cgcccgggta cccgtattcc
      661 caataaagcc tcttgctgtt tgcatccgaa tcgtggtctc gctgttcctt gggagggtct
      721 cctctgagtg attgactacc cacgacgggg gtctttcatt tgggggctcg tccgggattt
      781 ggagacccct gcccagggac caccgaccca ccaccgggag gtaagctggc cagcaactta
      841 tctgtgtctg tccgattgtc tagtgtctat gtttgatgtt atgcgcctgc gtctgtacta
      901 gttagctaac tagctctgta tctggcggac ccgtggtgga actgacgagt tctgaacacc
      961 cggccgcaac cctgggagac gtcccaggga ctttgggggc cgtttttgtg gcccgacctg
     1021 aggaagggag tcgatgtgga atccgacccc gtcaggatat gtggttctgg taggagacga
     1081 gaacctaaaa cagttcccgc ctccgtctga atttttgctt tcggtttgga accgaagccg
     1141 cgcgtcttgt ctgctgcagc gctgcagcat cgttctgtgt tgtctctgtc tgactgtgtt
     1201 tctgtatttg tctgaaaatt agggccagac tgttaccact cccttaagtt tgaccttagg
     1261 tcactggaaa gatgtcgagc ggatcgctca caaccagtcg gtagatgtca agaagagacg
     1321 ttgggttacc ttctgctctg cagaatggcc aacctttaac gtcggatggc cgcgagacgg
     1381 cacctttaac cgagacctca tcacccaggt taagatcaag gtcttttcac ctggcccgca
     1441 tggacaccca gaccaggtcc cctacatcgt gacctgggaa gccttggctt ttgacccccc
     1501 tccctgggtc aagccctttg tacaccctaa gcctccgcct cctcttcctc catccgcccc
     1561 gtctctcccc cttgaacctc ctcgttcgac cccgcctcga tcctcccttt atccagccct
     1621 cactccttct ctaggcgccg gaattgaaga tctgggggat cgatcctcta gagtccgtta
     1681 cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc ccattgacgt
     1741 caataatgac gtatgttccc atagtaacgc caatagggac tttccattga cgtcaatggg
     1801 tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat atgccaagta
     1861 cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc cagtacatga
     1921 ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct attaccatgg
     1981 tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca cggggatttc
     2041 caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat caacgggact
     2101 ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg cgtgtacggt
     2161 gggaggtcta tataagcaga gctcgtttag tgaaccgtca gatcgcctgg agacgccatc
     2221 cacgctgttt tgacctccat agaagacacc gggaccgatc cagcctccgc ggccgggaac
     2281 ggtgcattgg aacggacctg caggaattga tccgcggccg caccggggat caccatgggt
     2341 aagcctatcc ctaaccctct cctcggtctc gattctacgg gatctaccat ggtgagcgtg
     2401 atcaagcccg agatgaagat gaggtactac atggacggct ccgtcaatgg gcatgagttc
     2461 acaatcgagg gtgagggcac aggcagacct tacgagggac atcaggagat gacactgcgc
     2521 gtcacaatgg ccgagggcgg gccaatgcct ttcgccttcg acctggtgtc ccacgtgttc
     2581 tgttacggcc acagagtttt taccaagtac ccagaagaga tcccagacta tttcaagcag
     2641 gcctttcctg agggcctgtc ctgggagagg tccctggagt tcgaggacgg cggctccgcc
     2701 tccgtgagcg cccacatcag cctgaggggc aacaccttct accacaagtc caagttcacc
     2761 ggcgtgaact tccccgccga cggccccatc atgcagaacc agagcgtgga ctgggagccc
     2821 tccaccgaga agatcaccgc cagcgacggc gtgctgaagg gcgacgtgac catgtacctg
     2881 aagctggagg gcggcggcaa ccacaagtgc cagatgaaga ccacctacaa ggccgccaag
     2941 gagatcctgg agatgcccgg cgaccactac atcggccaca ggctggtgag gaagaccgag
     3001 ggcaacatca ccgagcaggt ggaggacgcc gtggcccact ccggtggcgg aggctcgggc
     3061 ggaggtgggt cgggatccgg ttctagagcc accggtatgt ctaatggtta tgaagaccac
     3121 atggccgaag actgcagggg tgacatcggg agaacgaatt tgatcgtcaa ctacctccct
     3181 cagaacatga cccaggatga gttacgaagc ctgttcagca gcattggtga agttgaatct
     3241 gcaaaactta ttcgggataa agtagcagga cacagcttgg gctatggctt tgtgaactac
     3301 gtgaccgcga aggatgcaga gagagcgatc aacacgctga acggcttgag gctccagtca
     3361 aaaaccatta aggtgtcgta tgctcgcccg agctcagagg tgatcaaaga cgccaacttg
     3421 tacatcagcg ggctcccgcg gaccatgacc cagaaggacg tagaagacat gttctctcgg
     3481 tttgggcgga tcatcaactc gcgggtcctc gtggatcaga ctacaggttt gtccagaggg
     3541 gttgcgttta tccggtttga caaacggtcg gaggcagaag aggcaattac cagtttcaat
     3601 ggtcataaac ccccaggttc ctctgagccc atcacagtga agtttgcagc caaccccaac
     3661 cagaacaaaa acgtggcact cctctcgcag ctgtaccact cgccagcgcg acggttcgga
     3721 ggccccgttc accaccaggc gcagagattc aggttctccc ccatgggcgt cgatcacatg
     3781 agcgggctct ctggcgtcaa cgtgccagga aacgcctcct ccggctggtg cattttcatc
     3841 tacaacctgg ggcaggatgc cgacgagggg atcctctggc agatgtttgg gccgtttggt
     3901 gccgtcacca atgtgaaagt gatccgcgac ttcaacacca acaagtgcaa agggtttggc
     3961 tttgtgacca tgacaaacta tgaagaagcc gcgatggcca tagccagcct gaacggctac
     4021 cgcctggggg acaaaatctt acaggtttcc ttcaaaacca acaagtccca caaataagaa
     4081 ttctgattaa ttccgccccc ctccctcccc cccccctaac gttactggcc gaagccgctt
     4141 ggaataaggc cggtgtgcgt ttgtctatat gttattttcc accatattgc cgtcttttgg
     4201 caatgtgagg gcccggaaac ctggccctgt cttcttgacg agcattccta ggggtctttc
     4261 ccctctcgcc aaaggaatgc aaggtctgtt gaatgtcgtg aaggaagcag ttcctctgga
     4321 agcttcttga agacaaacaa cgtctgtagc gaccctttgc aggcagcgga accccccacc
     4381 tggcgacagg tgcctctgcg gccaaaagcc acgtgtataa gatacacctg caaaggcggc
     4441 acaaccccag tgccacgttg tgagttggat agttgtggaa agagtcaaat ggctctcctc
     4501 aagcgtattc aacaaggggc tgaaggatgc ccagaaggta ccccattgta tgggatctga
     4561 tctggggcct cggtgcacat gctttacatg tgtttagtcg aggttaaaaa aacgtctagg
     4621 ccccccgaac cacggggacg tggttttcct ttgaaaaaca cgatgataag cttgatcacg
     4681 cgtctcgaga tgatccaccg gagcttacca tgaccgagta caagcccacg gtgcgcctcg
     4741 ccacccgcga cgacgtcccc agggccgtac gcaccctcgc cgccgcgttc gccgactacc
     4801 ccgccacgcg ccacaccgtc gatccggacc gccacatcga gcgggtcacc gagctgcaag
     4861 aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg
     4921 ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga
     4981 tcggcccgcg catggccgag ttgagcggtt cccggctggc cgcgcagcaa cagatggaag
     5041 gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc gtcggcgtct
     5101 cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt gctccccgga gtggaggcgg
     5161 ccgagcgcgc cggggtgccc gccttcctgg agacctccgc gccccgcaac ctccccttct
     5221 acgagcggct cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct
     5281 ggtgcatgac ccgcaagccc ggtgcctgac gcccgcccca cgacccgcag cgcccgaccg
     5341 aaaggagcgc acgaccccat gcatcgatca gtggtccagg ctctagtttt gactcaacaa
     5401 tatcaccagc tgaagcctat agagtacgag ccatagataa aataaaagat tttatttagt
     5461 ctccagaaaa aggggggaat gaaagacccc acctgtaggt ttggcaagct agcttaagta
     5521 acgccatttt gcaaggcatg gaaaaataca taactgagaa tagagaagtt cagatcaagg
     5581 tcaggaacag atggaacagg gtcgacccta gagaaccatc agatgtttcc agggtgcccc
     5641 aaggacctga aatgaccctg tgccttattt gaactaacca atcagttcgc ttctcgcttc
     5701 tgttcgcgcg cttctgctcc ccgagctcaa taaaagagcc cacaacccct cactcggggc
     5761 gccagtcctc cgattgactg agtcgcccgg gtacccgtgt atccaataaa ccctcttgca
     5821 gttgcatccg acttgtggtc tcgctgttcc ttgggagggt ctcctctgag tgattgacta
     5881 cccgtcagcg ggggtctttc atttgggggc tcgtccggga tcgggagacc cctgcccagg
     5941 gaccaccgac ccaccaccgg gaggtaagct ggctgcctcg cgcgtttcgg tgatgacggt
     6001 gaaaacctct gacacatgca gctcccggag acggtcacag cttgtctgta agcggatgcc
     6061 gggagcagac aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc
     6121 atgacccagt cacgtagcga tagcggagtg tagatccggc tgtggaatgt gtgtcagtta
     6181 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     6241 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     6301 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     6361 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     6421 gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     6481 ggcctaggct tttgcaaaaa gcttactggc ttaactatgc ggcatcagag cagattgtac
     6541 tgagagtgca ccatatgcgg tgtgaaatac cgcacagatg cgtaaggaga aaataccgca
     6601 tcaggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     6661 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     6721 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     6781 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     6841 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     6901 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     6961 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     7021 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     7081 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     7141 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     7201 agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga
     7261 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     7321 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     7381 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     7441 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     7501 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     7561 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     7621 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     7681 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     7741 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     7801 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     7861 ttgctgcagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     7921 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     7981 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     8041 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     8101 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     8161 cgtcaacacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     8221 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     8281 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     8341 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     8401 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     8461 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     8521 ttccccgaaa agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata
     8581 aaaataggcg tatcacgagg ccctttcgtc ttcaagaatt agcttggcca ttgcatacgt
     8641 tgtatccata tcataatatg tacatttata ttggctcatg tccaacatta ccgccatgtt
//
