LOCUS       sequence-226694-        8149 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Allows Gateway cloning of gene of interest into retroviral pMSCV 
            vector (with Neomycin/G418 resistance).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8149)
  AUTHORS   Harley ME, Murina O, Leitch A, Higgs MR, Bicknell LS, Yigit G, 
            Blackford AN, Zlatanou A, Mackenzie KJ, Reddy K, Halachev M, 
            McGlasson S, Reijns MAM, Fluteau A, Martin CA, Sabbioneda S, 
            Elcioglu NH, Altmuller J, Thiele H, Greenhalgh L, Chessa L, Maghnie 
            M, Salim M, Bober MB, Nurnberg P, Jackson SP, Hurles ME, Wollnik B, 
            Stewart GS, Jackson AP
  TITLE     TRAIP promotes DNA damage response during genome replication and is 
            mutated in primordial dwarfism.
  JOURNAL   Nat Genet. 2016 Jan;48(1):36-43. doi: 10.1038/ng.3451. Epub 2015 Nov
            23.
  PUBMED    26595769
REFERENCE   2  (bases 1 to 8149)
  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"; doi: "10.1038/ng.3451"; 
            journalName: "Nat Genet"; date: "2016-01"; volume: "48"; issue: "1";
            pages: "36-43"
COMMENT     Sequence Label: pMSCVneo-DEST
FEATURES             Location/Qualifiers
     source          1..8149
                     /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    1418..1542
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(1567..1572,1573..1590,1591..1597)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             1651..2310
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1717..1736)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             2652..2957
                     /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     2836..2855
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(2998..3122)
                     /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             3664..4467
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3727..3746)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4337..4356
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     LTR             4542..5056
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    5218..5234
                     /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(5242..5248,5249..5266,5267..5272))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5287..5308
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5425..5442)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5596..6184)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5676..5695)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6355..7146,7147..7215))
                     /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     6978..6997
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7216..7320)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7388..7406
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7444..7466)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7566..7585
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7779..7801
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7929..7948
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             join(8148..8149,1..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 gccggaattc gttatcaaca agtttgtaca aaaaagctga
     1441 acgagaaacg taaaatgata taaatatcaa tatattaaat tagattttgc ataaaaaaca
     1501 gactacataa tactgtaaaa cacaacatat ccagtcacta tggcggccgc attaggcacc
     1561 ccaggcttta cactttatgc ttccggctcg tataatgtgt ggattttgag ttaggatccg
     1621 tcgagatttt caggagctaa ggaagctaaa atggagaaaa aaatcactgg atataccacc
     1681 gttgatatat cccaatggca tcgtaaagaa cattttgagg catttcagtc agttgctcaa
     1741 tgtacctata accagaccgt tcagctggat attacggcct ttttaaagac cgtaaagaaa
     1801 aataagcaca agttttatcc ggcctttatt cacattcttg cccgcctgat gaatgctcat
     1861 ccggaattcc gtatggcaat gaaagacggt gagctggtga tatgggatag tgttcaccct
     1921 tgttacaccg ttttccatga gcaaactgaa acgttttcat cgctctggag tgaataccac
     1981 gacgatttcc ggcagtttct acacatatat tcgcaagatg tggcgtgtta cggtgaaaac
     2041 ctggcctatt tccctaaagg gtttattgag aatatgtttt tcgtctcagc caatccctgg
     2101 gtgagtttca ccagttttga tttaaacgtg gccaatatgg acaacttctt cgcccccgtt
     2161 ttcaccatgg gcaaatatta tacgcaaggc gacaaggtgc tgatgccgct ggcgattcag
     2221 gttcatcatg ccgtttgtga tggcttccat gtcggcagaa tgcttaatga attacaacag
     2281 tactgcgatg agtggcaggg cggggcgtaa agatctggat ccggcttact aaaagccaga
     2341 taacagtatg cgtatttgcg cgctgatttt tgcggtataa gaatatatac tgatatgtat
     2401 acccgaagta tgtcaaaaag aggtatgcta tgaagcagcg tattacagtg acagttgaca
     2461 gcgacagcta tcagttgctc aaggcatata tgatgtcaat atctccggtc tggtaagcac
     2521 aaccatgcag aatgaagccc gtcgtctgcg tgccgaacgc tggaaagcgg aaaatcagga
     2581 agggatggct gaggtcgccc ggtttattga aatgaacggc tcttttgctg acgagaacag
     2641 gggctggtga aatgcagttt aaggtttaca cctataaaag agagagccgt tatcgtctgt
     2701 ttgtggatgt acagagtgat attattgaca cgcccgggcg acggatggtg atccccctgg
     2761 ccagtgcacg tctgctgtca gataaagtct cccgtgaact ttacccggtg gtgcatatcg
     2821 gggatgaaag ctggcgcatg atgaccaccg atatggccag tgtgccggtc tccgttatcg
     2881 gggaagaagt ggctgatctc agccaccgcg aaaatgacat caaaaacgcc attaacctga
     2941 tgttctgggg aatataaatg tcaggctccc ttatacacag ccagtctgca ggtcgaccat
     3001 agtgactgga tatgttgtgt tttacagtat tatgtagtct gttttttatg caaaatctaa
     3061 tttaatatat tgatatttat atcattttac gtttctcgtt cagctttctt gtacaaagtg
     3121 gttgataacc tcgagagatc taattctacc 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 aatatgggat cggccattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg
     3721 gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg
     3781 ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc
     3841 ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct
     3901 tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa
     3961 gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg
     4021 gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa
     4081 gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat
     4141 gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg
     4201 cgcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc
     4261 atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac
     4321 cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg
     4381 gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc
     4441 tatcgccttc ttgacgagtt cttctgaggg gatccgtcga cctgcagcca agcttatcga
     4501 taaaataaaa gattttattt agtctccaga aaaagggggg aatgaaagac cccacctgta
     4561 ggtttggcaa gctagcttaa gtaacgccat tttgcaaggc atggaaaata cataactgag
     4621 aatagagaag ttcagatcaa ggttaggaac agagagacag cagaatatgg gccaaacagg
     4681 atatctgtgg taagcagttc ctgccccggc tcagggccaa gaacagatgg tccccagatg
     4741 cggtcccgcc ctcagcagtt tctagagaac catcagatgt ttccagggtg ccccaaggac
     4801 ctgaaatgac cctgtgcctt atttgaacta accaatcagt tcgcttctcg cttctgttcg
     4861 cgcgcttctg ctccccgagc tcaataaaag agcccacaac ccctcactcg gcgcgccagt
     4921 cctccgatag actgcgtcgc ccgggtaccc gtgtatccaa taaaccctct tgcagttgca
     4981 tccgacttgt ggtctcgctg ttccttggga gggtctcctc tgagtgattg actacccgtc
     5041 agcgggggtc tttcatgggt aacagtttct tgaagttgga gaacaacatt ctgagggtag
     5101 gagtcgaata ttaagtaatc ctgactcaat tagccactgt tttgaatcca catactccaa
     5161 tactcctgaa atagttcatt atggacagcg cagaaagagc tggggagaat tgtgaaattg
     5221 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg
     5281 tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc
     5341 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     5401 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     5461 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     5521 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     5581 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     5641 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     5701 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     5761 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     5821 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     5881 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     5941 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     6001 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     6061 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     6121 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     6181 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     6241 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     6301 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     6361 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     6421 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     6481 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     6541 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     6601 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     6661 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     6721 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     6781 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     6841 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     6901 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     6961 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     7021 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     7081 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     7141 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     7201 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     7261 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     7321 cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca tgacattaac
     7381 ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg cgtttcggtg atgacggtga
     7441 aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg
     7501 gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gctggcttaa
     7561 ctatgcggca tcagagcaga ttgtactgag agtgcaccat atgcggtgtg aaataccgca
     7621 cagatgcgta aggagaaaat accgcatcag gcgccattcg ccattcaggc tgcgcaactg
     7681 ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga aagggggatg
     7741 tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac gttgtaaaac
     7801 gacggcgcaa ggaatggtgc atgcaaggag atggcgccca acagtccccc ggccacgggg
     7861 cctgccacca tacccacgcc gaaacaagcg ctcatgagcc cgaagtggcg agcccgatct
     7921 tccccatcgg tgatgtcggc gatataggcg ccagcaaccg cacctgtggc gccggtgatg
     7981 ccggccacga tgcgtccggc gtagaggcga ttagtccaat ttgttaaaga caggatatca
     8041 gtggtccagg ctctagtttt gactcaacaa tatcaccagc tgaagcctat agagtacgag
     8101 ccatagataa aataaaagat tttatttagt ctccagaaaa aggggggaa
//
