LOCUS       sequence-207777-        8603 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  gamma-Retroviral transfer vector for expressing HuR (delHNS), V5 and
            mKO2 tags, IRES-driven Puromycin selection..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8603)
  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  JOURNAL   Unpublished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 8603)
  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            "Unpublished"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.DelHNS IRES Puro
FEATURES             Location/Qualifiers
     source          1..8603
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        310..513
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             514..689
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     misc_feature    752..951
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1152..1568
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1498..1520
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1536..1552
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     enhancer        1605..1908
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1909..2112
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2109..2133
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             2267..2308
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2318..2971
                     /codon_start=1
                     /product="monomeric Kusabira-Orange 2 fluorescent protein"
                     /label=mKO2
                     /translation="MVSVIKPEMKMRYYMDGSVNGHEFTIEGEGTGRPYEGHQEMTLRV
                     TMAEGGPMPFAFDLVSHVFCYGHRVFTKYPEEIPDYFKQAFPEGLSWERSLEFEDGGSA
                     SVSAHISLRGNTFYHKSKFTGVNFPADGPIMQNQSVDWEPSTEKITASDGVLKGDVTMY
                     LKLEGGGNHKCQMKTTYKAAKEILEMPGDHYIGHRLVRKTEGNITEQVEDAVAHS"
     CDS             2318..2971
                     /codon_start=1
                     /product="humanized monomeric Kusabira-Orange fluorescent 
                     protein"
                     /label=hmKO
                     /note="mammalian codon-optimized"
                     /translation="MVSVIKPEMKMRYYMDGSVNGHEFTIEGEGTGRPYEGHQEMTLRV
                     TMAEGGPMPFAFDLVSHVFCYGHRVFTKYPEEIPDYFKQAFPEGLSWERSLEFEDGGSA
                     SVSAHISLRGNTFYHKSKFTGVNFPADGPIMQNQSVDWEPSTEKITASDGVLKGDVTMY
                     LKLEGGGNHKCQMKTTYKAAKEILEMPGDHYIGHRLVRKTEGNITEQVEDAVAHS"
     misc_feature    3932..4504
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4099..4116)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     4326..4345
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4541..5140
                     /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(4541..4560)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5037..5057
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             5309..5734
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     Moloney murine leukemia virus"
     primer_bind     complement(5837..5859)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5995..6015)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6000..6329
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6180..6315
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6351..6370
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(6451..6468)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6622..7210)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6702..6721)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7381..8172,8173..8241))
                     /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     8004..8023
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8242..8346)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     8414..8432
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     enhancer        join(8532..8603,1..309)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc
       61 cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac
      121 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
      181 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgctctg gcattatgcc
      241 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
      301 attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca
      361 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat
      421 caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg
      481 cgtgtacggt gggaggtcta tataagcaga gctcaataaa agagcccaca acccctcact
      541 cggcgcgcca gtcttccgat agactgcgtc gcccgggtac ccgtattccc aataaagcct
      601 cttgctgttt gcatccgaat cgtggtctcg ctgttccttg ggagggtctc ctctgagtga
      661 ttgactaccc acgacggggg tctttcattt gggggctcgt ccgggatttg gagacccctg
      721 cccagggacc accgacccac caccgggagg taagctggcc agcaacttat ctgtgtctgt
      781 ccgattgtct agtgtctatg tttgatgtta tgcgcctgcg tctgtactag ttagctaact
      841 agctctgtat ctggcggacc cgtggtggaa ctgacgagtt ctgaacaccc ggccgcaacc
      901 ctgggagacg tcccagggac tttgggggcc gtttttgtgg cccgacctga ggaagggagt
      961 cgatgtggaa tccgaccccg tcaggatatg tggttctggt aggagacgag aacctaaaac
     1021 agttcccgcc tccgtctgaa tttttgcttt cggtttggaa ccgaagccgc gcgtcttgtc
     1081 tgctgcagcg ctgcagcatc gttctgtgtt gtctctgtct gactgtgttt ctgtatttgt
     1141 ctgaaaatta gggccagact gttaccactc ccttaagttt gaccttaggt cactggaaag
     1201 atgtcgagcg gatcgctcac aaccagtcgg tagatgtcaa gaagagacgt tgggttacct
     1261 tctgctctgc agaatggcca acctttaacg tcggatggcc gcgagacggc acctttaacc
     1321 gagacctcat cacccaggtt aagatcaagg tcttttcacc tggcccgcat ggacacccag
     1381 accaggtccc ctacatcgtg acctgggaag ccttggcttt tgacccccct ccctgggtca
     1441 agccctttgt acaccctaag cctccgcctc ctcttcctcc atccgccccg tctctccccc
     1501 ttgaacctcc tcgttcgacc ccgcctcgat cctcccttta tccagccctc actccttctc
     1561 taggcgccgg aattgaagat ctgggggatc gatcctctag agtccgttac ataacttacg
     1621 gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg
     1681 tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta
     1741 cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt
     1801 gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac
     1861 tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt gatgcggttt
     1921 tggcagtaca tcaatgggcg tgaatagcgg tttgactcac ggggatttcc aagtctccac
     1981 cccattgacg tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt
     2041 cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat
     2101 ataagcagag ctcgtttagt gaaccgtcag atcgcctgga gacgccatcc acgctgtttt
     2161 gacctccata gaagacaccg ggaccgatcc agcctccgcg gccgggaacg gtgcattgga
     2221 acggacctgc aggaattgat ccgcggccgc accggggatc accatgggta agcctatccc
     2281 taaccctctc ctcggtctcg attctacggg atctaccatg gtgagcgtga tcaagcccga
     2341 gatgaagatg aggtactaca tggacggctc cgtcaatggg catgagttca caatcgaggg
     2401 tgagggcaca ggcagacctt acgagggaca tcaggagatg acactgcgcg tcacaatggc
     2461 cgagggcggg ccaatgcctt tcgccttcga cctggtgtcc cacgtgttct gttacggcca
     2521 cagagttttt accaagtacc cagaagagat cccagactat ttcaagcagg cctttcctga
     2581 gggcctgtcc tgggagaggt ccctggagtt cgaggacggc ggctccgcct ccgtgagcgc
     2641 ccacatcagc ctgaggggca acaccttcta ccacaagtcc aagttcaccg gcgtgaactt
     2701 ccccgccgac ggccccatca tgcagaacca gagcgtggac tgggagccct ccaccgagaa
     2761 gatcaccgcc agcgacggcg tgctgaaggg cgacgtgacc atgtacctga agctggaggg
     2821 cggcggcaac cacaagtgcc agatgaagac cacctacaag gccgccaagg agatcctgga
     2881 gatgcccggc gaccactaca tcggccacag gctggtgagg aagaccgagg gcaacatcac
     2941 cgagcaggtg gaggacgccg tggcccactc cggtggcgga ggctcgggcg gaggtgggtc
     3001 gggatccggt tctagagcca ccggtatgtc taatggttat gaagaccaca tggccgaaga
     3061 ctgcaggggt gacatcggga gaacgaattt gatcgtcaac tacctccctc agaacatgac
     3121 ccaggatgag ttacgaagcc tgttcagcag cattggtgaa gttgaatctg caaaacttat
     3181 tcgggataaa gtagcaggac acagcttggg ctatggcttt gtgaactacg tgaccgcgaa
     3241 ggatgcagag agagcgatca acacgctgaa cggcttgagg ctccagtcaa aaaccattaa
     3301 ggtgtcgtat gctcgcccga gctcagaggt gatcaaagac gccaacttgt acatcagcgg
     3361 gctcccgcgg accatgaccc agaagtacgt agaagacatg ttctctcggt ttgggcggat
     3421 catcaactcg cgggtcctcg tggatcagac tacaggtttg tccagagggg ttgcgtttat
     3481 ccggtttgac aaacggtcgg aggcagaaga ggcaattacc agtttcaatg gtcataaacc
     3541 cccaggttcc tctgagccca tcacagtgaa gtttgcagcc aaccccaacc agaacaaaaa
     3601 cgtggcactc ctctcgcagc tgtaccactc gccagcgaag cttggaaacg cctcctccgg
     3661 ctggtgcatt ttcatctaca acctggggca ggatgccgac gaggggatcc tctggcagat
     3721 gtttgggccg tttggtgccg tcaccaatgt gaaagtgatc cgcgacttca acaccaacaa
     3781 gtgcaaaggg tttggctttg tgaccatgac aaactatgaa gaagccgcga tggccatagc
     3841 cagcctgaac ggctaccgcc tgggggacaa aatcttacag gtttccttca aaaccaacaa
     3901 gtcccacaaa taagaattct gattaattcc gcccctctcc ctcccccccc cctaacgtta
     3961 ctggccgaag ccgcttggaa taaggccggt gtgcgtttgt ctatatgtta ttttccacca
     4021 tattgccgtc ttttggcaat gtgagggccc ggaaacctgg ccctgtcttc ttgacgagca
     4081 ttcctagggg tctttcccct ctcgccaaag gaatgcaagg tctgttgaat gtcgtgaagg
     4141 aagcagttcc tctggaagct tcttgaagac aaacaacgtc tgtagcgacc ctttgcaggc
     4201 agcggaaccc cccacctggc gacaggtgcc tctgcggcca aaagccacgt gtataagata
     4261 cacctgcaaa ggcggcacaa ccccagtgcc acgttgtgag ttggatagtt gtggaaagag
     4321 tcaaatggct ctcctcaagc gtattcaaca aggggctgaa ggatgcccag aaggtacccc
     4381 attgtatggg atctgatctg gggcctcggt gcacatgctt tacatgtgtt tagtcgaggt
     4441 taaaaaacgt ctaggccccc cgaaccacgg ggacgtggtt ttcctttgaa aaacacgatg
     4501 ataagcttga tcacgcgtct cgagatcacc ggagcttacc atgaccgagt acaagcccac
     4561 ggtgcgcctc gccacccgcg acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt
     4621 cgccgactac cccgccacgc gccacaccgt cgatccggac cgccacatcg agcgggtcac
     4681 cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc
     4741 ggacgacggc gccgcggtgg cggtctggac cacgccggag agcgtcgaag cgggggcggt
     4801 gttcgccgag atcggcccgc gcatggccga gttgagcggt tcccggctgg ccgcgcagca
     4861 acagatggaa ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt tcctggccac
     4921 cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc agcgccgtcg tgctccccgg
     4981 agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg gagacctccg cgccccgcaa
     5041 cctccccttc tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg
     5101 accgcgcacc tggtgcatga cccgcaagcc cggtgcctga cgcccgcccc acgacccgca
     5161 gcgcccgacc gaaaggagcg cacgacccca tgcatcgatc agtggtccag gctctagttt
     5221 tgactcaaca atatcaccag ctgaagccta tagagtacga gccatagata aaataaaaga
     5281 ttttatttag tctccagaaa aaggggggaa tgaaagaccc cacctgtagg tttggcaagc
     5341 tagcttaagt aacgccattt tgcaaggcat ggaaaaatac ataactgaga atagagaagt
     5401 tcagatcaag gtcaggaaca gatggaacag ggtcgaccct agagaaccat cagatgtttc
     5461 cagggtgccc caaggacctg aaatgaccct gtgccttatt tgaactaacc aatcagttcg
     5521 cttctcgctt ctgttcgcgc gcttctgctc cccgagctca ataaaagagc ccacaacccc
     5581 tcactcgggg cgccagtcct ccgattgact gagtcgcccg ggtacccgtg tatccaataa
     5641 accctcttgc agttgcatcc gacttgtggt ctcgctgttc cttgggaggg tctcctctga
     5701 gtgattgact acccgtcagc gggggtcttt catttggggg ctcgtccggg atcgggagac
     5761 ccctgcccag ggaccaccga cccaccaccg ggaggtaagc tggctgcctc gcgcgtttcg
     5821 gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt
     5881 aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc
     5941 ggggcgcagc catgacccag tcacgtagcg atagcggagt gtagatccgg ctgtggaatg
     6001 tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     6061 tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa
     6121 gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta actccgccca
     6181 tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt
     6241 ttatttatgc agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag
     6301 gcttttttgg aggcctaggc ttttgcaaaa agcttactgg cttaactatg cggcatcaga
     6361 gcagattgta ctgagagtgc accatatgcg gtgtgaaata ccgcacagat gcgtaaggag
     6421 aaaataccgc atcaggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     6481 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     6541 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     6601 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     6661 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     6721 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     6781 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     6841 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     6901 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     6961 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     7021 agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat ttggtatctg
     7081 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     7141 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     7201 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     7261 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     7321 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     7381 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     7441 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     7501 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     7561 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     7621 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     7681 cgttgttgcc attgctgcag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     7741 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     7801 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     7861 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     7921 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     7981 ctcttgcccg gcgtcaacac gggataatac cgcgccacat agcagaactt taaaagtgct
     8041 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     8101 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     8161 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     8221 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     8281 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     8341 tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac
     8401 attaacctat aaaaataggc gtatcacgag gccctttcgt cttcaagaat tagcttggcc
     8461 attgcatacg ttgtatccat atcataatat gtacatttat attggctcat gtccaacatt
     8521 accgccatgt tgacattgat tattgactag ttattaatag taatcaatta cggggtcatt
     8581 agttcatagc ccatatatgg agt
//
