LOCUS       sequence-226693-        7977 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Allows Gateway cloning of gene of interest into retroviral pMSCV 
            vector (with Puromycin resistance).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7977)
  AUTHORS   Benitez-Guijarro M, Lopez-Ruiz C, Tarnauskaite Z, Murina O, Mian 
            Mohammad M, Williams TC, Fluteau A, Sanchez L, Vilar-Astasio R, 
            Garcia-Canadas M, Cano D, Kempen MH, Sanchez-Pozo A, Heras SR, 
            Jackson AP, Reijns MA, Garcia-Perez JL
  TITLE     RNase H2, mutated in Aicardi-Goutieres syndrome, promotes LINE-1 
            retrotransposition.
  JOURNAL   EMBO J. 2018 Aug 1;37(15). pii: embj.201798506. doi: 
            10.15252/embj.201798506. Epub 2018 Jun 29.
  PUBMED    29959219
REFERENCE   2  (bases 1 to 7977)
  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: "EMBO J."; 
            date: "2018-08-1"; volume: "37(15). pii: embj.201798506. doi"; 
            pages: " 10.15252/embj.201798506. Epub 2018 Jun 29"
COMMENT     Sequence Label: pMSCVpuro-DEST
FEATURES             Location/Qualifiers
     source          1..7977
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    579..920
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             987..1403
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1333..1355
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1371..1387
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     protein_bind    1429..1553
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(1578..1583,1584..1601,1602..1608)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             1662..2321
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1728..1747)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             2663..2968
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     2847..2866
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(3009..3133)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        3151..3650
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(3186..3209)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     3583..3602
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             3671..4270
                     /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(3671..3690)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4167..4187
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             4370..4884
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    5046..5062
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(5070..5076,5077..5094,5095..5100))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5115..5136
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5253..5270)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5424..6012)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5504..5523)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6183..6974,6975..7043))
                     /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     6806..6825
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7044..7148)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7216..7234
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7272..7294)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7394..7413
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7607..7629
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7757..7776
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             join(7976..7977,1..515)
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
ORIGIN
        1 tgaaagaccc cacctgtagg tttggcaagc tagcttaagt aacgccattt tgcaaggcat
       61 ggaaaataca taactgagaa tagagaagtt cagatcaagg ttaggaacag agagacagca
      121 gaatatgggc caaacaggat atctgtggta agcagttcct gccccggctc agggccaaga
      181 acagatggtc cccagatgcg gtcccgccct cagcagtttc tagagaacca tcagatgttt
      241 ccagggtgcc ccaaggacct gaaatgaccc tgtgccttat ttgaactaac caatcagttc
      301 gcttctcgct tctgttcgcg cgcttctgct ccccgagctc aataaaagag cccacaaccc
      361 ctcactcggc gcgccagtcc tccgatagac tgcgtcgccc gggtacccgt attcccaata
      421 aagcctcttg ctgtttgcat ccgaatcgtg gactcgctga tccttgggag ggtctcctca
      481 gattgattga ctgcccacct cgggggtctt tcatttggag gttccaccga gatttggaga
      541 cccctgccca gggaccaccg acccccccgc cgggaggtaa gctggccagc ggtcgtttcg
      601 tgtctgtctc tgtctttgtg cgtgtttgtg ccggcatcta atgtttgcgc ctgcgtctgt
      661 actagttagc taactagctc tgtatctggc ggacccgtgg tggaactgac gagttctgaa
      721 cacccggccg caaccctggg agacgtccca gggactttgg gggccgtttt tgtggcccga
      781 cctgaggaag ggagtcgatg tggaatccga ccccgtcagg atatgtggtt ctggtaggag
      841 acgagaacct aaaacagttc ccgcctccgt ctgaattttt gctttcggtt tggaaccgaa
      901 gccgcgcgtc ttgtctgctg cagcgctgca gcatcgttct gtgttgtctc tgtctgactg
      961 tgtttctgta tttgtctgaa aattagggcc agactgttac cactccctta agtttgacct
     1021 taggtcactg gaaagatgtc gagcggatcg ctcacaacca gtcggtagat gtcaagaaga
     1081 gacgttgggt taccttctgc tctgcagaat ggccaacctt taacgtcgga tggccgcgag
     1141 acggcacctt taaccgagac ctcatcaccc aggttaagat caaggtcttt tcacctggcc
     1201 cgcatggaca cccagaccag gtcccctaca tcgtgacctg ggaagccttg gcttttgacc
     1261 cccctccctg ggtcaagccc tttgtacacc ctaagcctcc gcctcctctt cctccatccg
     1321 ccccgtctct cccccttgaa cctcctcgtt cgaccccgcc tcgatcctcc ctttatccag
     1381 ccctcactcc ttctctaggc gccggaatta gatctctcga ggttatcaac aagtttgtac
     1441 aaaaaagctg aacgagaaac gtaaaatgat ataaatatca atatattaaa ttagattttg
     1501 cataaaaaac agactacata atactgtaaa acacaacata tccagtcact atggcggccg
     1561 cattaggcac cccaggcttt acactttatg cttccggctc gtataatgtg tggattttga
     1621 gttaggatcc gtcgagattt tcaggagcta aggaagctaa aatggagaaa aaaatcactg
     1681 gatataccac cgttgatata tcccaatggc atcgtaaaga acattttgag gcatttcagt
     1741 cagttgctca atgtacctat aaccagaccg ttcagctgga tattacggcc tttttaaaga
     1801 ccgtaaagaa aaataagcac aagttttatc cggcctttat tcacattctt gcccgcctga
     1861 tgaatgctca tccggaattc cgtatggcaa tgaaagacgg tgagctggtg atatgggata
     1921 gtgttcaccc ttgttacacc gttttccatg agcaaactga aacgttttca tcgctctgga
     1981 gtgaatacca cgacgatttc cggcagtttc tacacatata ttcgcaagat gtggcgtgtt
     2041 acggtgaaaa cctggcctat ttccctaaag ggtttattga gaatatgttt ttcgtctcag
     2101 ccaatccctg ggtgagtttc accagttttg atttaaacgt ggccaatatg gacaacttct
     2161 tcgcccccgt tttcaccatg ggcaaatatt atacgcaagg cgacaaggtg ctgatgccgc
     2221 tggcgattca ggttcatcat gccgtttgtg atggcttcca tgtcggcaga atgcttaatg
     2281 aattacaaca gtactgcgat gagtggcagg gcggggcgta aagatctgga tccggcttac
     2341 taaaagccag ataacagtat gcgtatttgc gcgctgattt ttgcggtata agaatatata
     2401 ctgatatgta tacccgaagt atgtcaaaaa gaggtatgct atgaagcagc gtattacagt
     2461 gacagttgac agcgacagct atcagttgct caaggcatat atgatgtcaa tatctccggt
     2521 ctggtaagca caaccatgca gaatgaagcc cgtcgtctgc gtgccgaacg ctggaaagcg
     2581 gaaaatcagg aagggatggc tgaggtcgcc cggtttattg aaatgaacgg ctcttttgct
     2641 gacgagaaca ggggctggtg aaatgcagtt taaggtttac acctataaaa gagagagccg
     2701 ttatcgtctg tttgtggatg tacagagtga tattattgac acgcccgggc gacggatggt
     2761 gatccccctg gccagtgcac gtctgctgtc agataaagtc tcccgtgaac tttacccggt
     2821 ggtgcatatc ggggatgaaa gctggcgcat gatgaccacc gatatggcca gtgtgccggt
     2881 ctccgttatc ggggaagaag tggctgatct cagccaccgc gaaaatgaca tcaaaaacgc
     2941 cattaacctg atgttctggg gaatataaat gtcaggctcc cttatacaca gccagtctgc
     3001 aggtcgacca tagtgactgg atatgttgtg ttttacagta ttatgtagtc tgttttttat
     3061 gcaaaatcta atttaatata ttgatattta tatcatttta cgtttctcgt tcagctttct
     3121 tgtacaaagt ggttgataac gaattctacc gggtagggga ggcgcttttc ccaaggcagt
     3181 ctggagcatg cgctttagca gccccgctgg gcacttggcg ctacacaagt ggcctctggc
     3241 ctcgcacaca ttccacatcc accggtaggc gccaaccggc tccgttcttt ggtggcccct
     3301 tcgcgccacc ttctactcct cccctagtca ggaagttccc ccccgccccg cagctcgcgt
     3361 cgtgcaggac gtgacaaatg gaagtagcac gtctcactag tctcgtgcag atggacagca
     3421 ccgctgagca atggaagcgg gtaggccttt ggggcagcgg ccaatagcag ctttgctcct
     3481 tcgctttctg ggctcagagg ctgggaaggg gtgggtccgg gggcgggctc aggggcgggc
     3541 tcaggggcgg ggcgggcgcc cgaaggtcct ccggaggccc ggcattctgc acgcttcaaa
     3601 agcgcacgtc tgccgcgctg ttctcctctt cctcatctcc gggcctttcg acctgcagcc
     3661 caagcttacc atgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc
     3721 cagggccgta cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt
     3781 cgatccggac cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt
     3841 cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac
     3901 cacgccggag agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga
     3961 gttgagcggt tcccggctgg ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg
     4021 gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc accagggcaa
     4081 gggtctgggc agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc
     4141 cgccttcctg gagacctccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac
     4201 cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc
     4261 cggtgcctga cgcccgcccc acgacccgca gcgcccgacc gaaaggagcg cacgacccca
     4321 tgcatcgata aaataaaaga ttttatttag tctccagaaa aaggggggaa tgaaagaccc
     4381 cacctgtagg tttggcaagc tagcttaagt aacgccattt tgcaaggcat ggaaaataca
     4441 taactgagaa tagagaagtt cagatcaagg ttaggaacag agagacagca gaatatgggc
     4501 caaacaggat atctgtggta agcagttcct gccccggctc agggccaaga acagatggtc
     4561 cccagatgcg gtcccgccct cagcagtttc tagagaacca tcagatgttt ccagggtgcc
     4621 ccaaggacct gaaatgaccc tgtgccttat ttgaactaac caatcagttc gcttctcgct
     4681 tctgttcgcg cgcttctgct ccccgagctc aataaaagag cccacaaccc ctcactcggc
     4741 gcgccagtcc tccgatagac tgcgtcgccc gggtacccgt gtatccaata aaccctcttg
     4801 cagttgcatc cgacttgtgg tctcgctgtt ccttgggagg gtctcctctg agtgattgac
     4861 tacccgtcag cgggggtctt tcatgggtaa cagtttcttg aagttggaga acaacattct
     4921 gagggtagga gtcgaatatt aagtaatcct gactcaatta gccactgttt tgaatccaca
     4981 tactccaata ctcctgaaat agttcattat ggacagcgca gaaagagctg gggagaattg
     5041 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     5101 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     5161 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     5221 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     5281 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     5341 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     5401 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     5461 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     5521 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     5581 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     5641 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     5701 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     5761 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     5821 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     5881 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     5941 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     6001 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     6061 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     6121 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     6181 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     6241 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     6301 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     6361 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     6421 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     6481 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     6541 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     6601 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     6661 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     6721 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     6781 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     6841 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     6901 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     6961 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     7021 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     7081 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     7141 gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg
     7201 acattaacct ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat
     7261 gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg
     7321 gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc
     7381 tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccatat gcggtgtgaa
     7441 ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc gccattcgcc attcaggctg
     7501 cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa
     7561 gggggatgtg ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt
     7621 tgtaaaacga cggcgcaagg aatggtgcat gcaaggagat ggcgcccaac agtcccccgg
     7681 ccacggggcc tgccaccata cccacgccga aacaagcgct catgagcccg aagtggcgag
     7741 cccgatcttc cccatcggtg atgtcggcga tataggcgcc agcaaccgca cctgtggcgc
     7801 cggtgatgcc ggccacgatg cgtccggcgt agaggcgatt agtccaattt gttaaagaca
     7861 ggatatcagt ggtccaggct ctagttttga ctcaacaata tcaccagctg aagcctatag
     7921 agtacgagcc atagataaaa taaaagattt tatttagtct ccagaaaaag gggggaa
//
