LOCUS       sequence-270083-        8366 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  All-in-one Sleeping Beauty vector for stable transfection of 
            eukaryotic cells with hygromycin restistance. CMV protomor drives 
            SB100X transposase for transposon integration..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8366)
  AUTHORS   Flies AS, Darby JM, Lennard PR, Murphy PR, Ong CEB, Pinfold TL, De 
            Luca A, Lyons AB, Woods GM, Patchett AL
  TITLE     A novel system to map protein interactions reveals evolutionarily 
            conserved immune evasion pathways on transmissible cancers.
  JOURNAL   Sci Adv. 2020 Jul 1;6(27). pii: 6/27/eaba5031. doi: 
            10.1126/sciadv.aba5031. Print 2020 Jul.
  PUBMED    32937435
REFERENCE   2  (bases 1 to 8366)
  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: "Sci Adv."; 
            date: "2020-07-1"; volume: "6(27). pii: 6/27/eaba5031. doi"; pages: 
            " 10.1126/sciadv.aba5031. Print 2020 Jul"
COMMENT     Sequence Label: pAF123
FEATURES             Location/Qualifiers
     source          1..8366
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      26..35
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(275..292)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    281..505
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     enhancer        867..1170
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1171..1374
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1325..1345
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1371..1395
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          1512..1608
                     /label=SV40 intron
                     /note="modified SV40 intron with splice donor and acceptor 
                     sites"
     primer_bind     1665..1684
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1665..1683
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1694..2716
                     /codon_start=1
                     /product="hyperactive variant of the Sleeping Beauty 
                     transposase (Mates et al., 2009)"
                     /label=SB100X
                     /translation="MGKSKEISQDLRKRIVDLHKSGSSLGAISKRLAVPRSSVQTIVRK
                     YKHHGTTQPSYRSGRRRVLSPRDERTLVRKVQINPRTTAKDLVKMLEETGTKVSISTVK
                     RVLYRHNLKGHSARKKPLLQNRHKKARLRFATAHGDKDRTFWRNVLWSDETKIELFGHN
                     DHRYVWRKKGEACKPKNTIPTVKHGGGSIMLWGCFAAGGTGALHKIDGIMDAVQYVDIL
                     KQHLKTSVRKLKLGRKWVFQHDNDPKHTSKVVAKWLKDNKVKVLEWPSQSPDLNPIENL
                     WAELKKRVRARRPTNLTQLHQLCQEEWAKIHPNYCGKLVEGYPKRLTQVKQFKGNATKY
                     "
     polyA_signal    2733..2867
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2757..2776)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2811..2830
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3020..3124
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3125..3193,3194..3985)
                     /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(3343..3362)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4156..4744
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4645..4664
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4898..4915
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     5289..5308
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     polyA_signal    5307..5362
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     5408..5427
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     CDS             complement(5527..6552)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             complement(5527..6552)
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     promoter        7203..7414
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     primer_bind     8314..8334
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
ORIGIN
        1 aaactacccc aagctggcct ctgaggccac catggctcga gtttcgaact caattatact
       61 gaattgtttc atcatagggt tcaacattgt gataaatcag gacttcaatg tctagtgttc
      121 agtaaatacc ttgtagggct ctccaaaaag gggaatcttt tcaggaaatg cctctttctc
      181 ttggtgggct tcctggttgc gtctttcctt ctcactaatt cgaaggcctg tcaggccaag
      241 cttggggatc aattctctag agctcgctga tcagcctcga ctgtgccttc tagttgccag
      301 ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact
      361 gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt
      421 ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat
      481 gctggggatg cggtgggctc tatggcttct gaggcggaaa gaaccagctg ggcctactag
      541 atagcggacc ccttaccgaa acatcgccgc attctgcaga ggagtcgaac ttagtgtatg
      601 taaacttctg acccactgga attgtgatac agtgaattat aagtgaaata atctgtctgt
      661 aaacaattgt tggaaaaatg acttgtgtca tgcacaaagt agatgtccta actgacttgc
      721 caaaactatt gtttgttaac aagaaatttg tggagtagtt gaaaaacgag ttttaatgac
      781 tccaacttaa gtgtatgtaa acttccgact tcaactgtat agggatccaa ttttaacaaa
      841 atattcgagc ttgcatgcct gcaggtcgtt acataactta cggtaaatgg cccgcctggc
      901 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
      961 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
     1021 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
     1081 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
     1141 atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg
     1201 cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg
     1261 agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca
     1321 ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta
     1381 gtgaaccgtc agatcgcctg gagacgccat ccacgctgtt ttgacctcca tagaagacac
     1441 cgggaccgat ccagcctccg gactctagag gatccggtac tcgaggaact gaaaaaccag
     1501 aaagttaact ggtaagttta gtctttttgt cttttatttc aggtcccgga tccggtggtg
     1561 gtgcaaatca aagaactgct cctcagtgga tgttgccttt acttctaggc ctgtacggaa
     1621 gtgttacttc tgctctaaaa gctgcggaat tgtacccgcg gaattaatac gactcactat
     1681 agggactagt accatgggaa aatcaaaaga aatcagccaa gacctcagaa aaagaattgt
     1741 agacctccac aagtctggtt catccttggg agcaatttcc aaacgcctgg cggtaccacg
     1801 ttcatctgta caaacaatag tacgcaagta taaacaccat gggaccacgc agccgtcata
     1861 ccgctcagga aggagacgcg ttctgtctcc tagagatgaa cgtactttgg tgcgaaaagt
     1921 gcaaatcaat cccagaacaa cagcaaagga ccttgtgaag atgctggagg aaacaggtac
     1981 aaaagtatct atatccacag taaaacgagt cctatatcga cataacctga aaggccactc
     2041 agcaaggaag aagccactgc tccaaaaccg acataagaaa gccagactac ggtttgcaac
     2101 tgcacatggg gacaaagatc gtactttttg gagaaatgtc ctctggtctg atgaaacaaa
     2161 aatagaactg tttggccata atgaccatcg ttatgtttgg aggaagaagg gggaggcttg
     2221 caagccgaag aacaccatcc caaccgtgaa gcacgggggt ggcagcatca tgttgtgggg
     2281 gtgctttgct gcaggaggga ctggtgcact tcacaaaata gatggcatca tggacgcggt
     2341 gcagtatgtg gatatattga agcaacatct caagacatca gtcaggaagt taaagcttgg
     2401 tcgcaaatgg gtcttccaac acgacaatga ccccaagcat acttccaaag ttgtggcaaa
     2461 atggcttaag gacaacaaag tcaaggtatt ggagtggcca tcacaaagcc ctgacctcaa
     2521 tcctatagaa aatttgtggg cagaactgaa aaagcgtgtg cgagcaagga ggcctacaaa
     2581 cctgactcag ttacaccagc tctgtcagga ggaatgggcc aaaattcacc caaattattg
     2641 tgggaagctt gtggaaggct acccgaaacg tttgacccaa gttaaacaat ttaaaggcaa
     2701 tgctaccaaa tactaggggc cctaaccgcg gggatccaga catgataaga tacattgatg
     2761 agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg
     2821 atgctattgc tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt
     2881 gcattcattt tatgtttcag gttcaggggg aggtgtggga ggttttttaa cgcttacaat
     2941 ttcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg catcaggtgg
     3001 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     3061 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     3121 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     3181 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     3241 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     3301 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     3361 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     3421 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     3481 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     3541 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     3601 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     3661 gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct
     3721 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
     3781 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
     3841 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
     3901 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     3961 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat
     4021 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
     4081 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     4141 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     4201 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     4261 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
     4321 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     4381 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     4441 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     4501 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     4561 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     4621 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     4681 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     4741 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     4801 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
     4861 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
     4921 ggatccctat acagttgaag tcggaagttt acatacacct tagccaaata catttaaact
     4981 cactttttca caattcctga catttaatcc tagtaaaaat tccctgtctt aggtcagtta
     5041 ggatcaccac tttattttaa gaatgtgaaa tatcagaata atagtagaga gaatgattca
     5101 tttcagcttt tatttctttc atcacattcc cagtgggtca gaagtttaca tacactcgac
     5161 tcctctgcag aatgcggcga tgtttcggta aggggtcccg agtagctagt tcatggcagc
     5221 cagcatatgg catatgttgc caaactctaa accaaatact cattctgatg ttttaaatga
     5281 tttgccctcc catatgtcct tccgagtgag agacacaaaa aattccaaca cactattgca
     5341 atgaaaataa atttccttta ttagccagaa gtcagatgct caaggggctt catgatgtcc
     5401 ccataatttt tggcagaggg aaaaagatct cagtggtatt tgtgagccag ggcattggcc
     5461 acaccagcca ccaccttctg ataggcagcc tgcacctgag gagtgaattc acgacaggcc
     5521 ttcgaactat tcctttgccc tcggacgagt gctggggcgt cggtttccac tatcggcgag
     5581 tacttctaca cagccatcgg tccagacggc cgcgcttctg cgggcgattt gtgtacgccc
     5641 gacagtcccg gctccggatc ggacgattgc gtcgcatcga ccctgcgccc aagctgcatc
     5701 atcgaaattg ccgtcaacca agctctgata gagttggtca agaccaatgc ggagcatata
     5761 cgcccggagc cgcggcgatc ctgcaagctc cggatgcctc cgctcgaagt agcgcgtctg
     5821 ctgctccata caagccaacc acggcctcca gaagaagatg ttggcgacct cgtattggga
     5881 atccccgaac atcgcctcgc tccagtcaat gaccgctgtt atgcggccat tgtccgtcag
     5941 gacattgttg gagccgaaat ccgcgtgcac gaggtgccgg acttcggggc agtcctcggc
     6001 ccaaagcatc agctcatcga gagcctgcgc gacggacgca ctgacggtgt cgtccatcac
     6061 agtttgccag tgatacacat ggggatcagc aatcgcgcat atgaaatcac gccatgtagt
     6121 gtattgaccg attccttgcg gtccgaatgg gccgaacccg ctcgtctggc taagatcggc
     6181 cgcagcgatc gcatccatgg cctccgcgac cggctgcaga acagcgggca gttcggtttc
     6241 aggcaggtct tgcaacgtga caccctgtgc acggcgggag atgcaatagg tcaggctctc
     6301 gctgaattcc ccaatgtcaa gcacttccgg aatcgggagc gcggccgatg caaagtgccg
     6361 ataaacataa cgatctttgt agaaaccatc ggcgcagcta tttacccgca ggacatatcc
     6421 acgccctcct acatcgaagc tgaaagcacg agattcttcg ccctccgaga gctgcatcag
     6481 gtcggagacg ctgtcgaact tttcgatcag aaacttctcg acagacgtcg cggtgagttc
     6541 aggctttttc atggtggcct caggtgcaga ggtttctaca gggaaagaga gaaaggctac
     6601 agttagtttg ctgtgatgat gctaagtgcc gaggtcagat gtctatggat tgtacacgga
     6661 atcccaacgc aggaaccaaa aaacactcac ccaaaaaaat ggcgctaagg ccgagagaaa
     6721 ggcgaagttc cgtctacggc tatatacctt gaaacagggg cggcagaagt gacgcaacgc
     6781 ggttgcgcgg acggcttcct gtgttttcat tggtcgaagc gttctgcgcg tgcgcagaga
     6841 ttccccaggg aaggcgggcc ctcttccgaa tgtccaatcc ggacattctt ccggttggac
     6901 ctccgcggtc agcgctgtgc ccggcgagag atcacgtggg gcgcggaggc ggtgctgctg
     6961 gggcacggcc gtccagcctc ggcggccata tttttgaggg gctgttcatc tcgttcacac
     7021 gctctgtccg ccatgtttgt gagtggaagc gccattaccc cttcaagcga ctgaaggctg
     7081 cagggcctct ggtggcccgc atggggagac cagacccgcc aggcccgcct ttccgcactc
     7141 agtccgggct tactttattt tgtgagacag ggtctcgcct agaggctccg gtgcccgtca
     7201 gtgggcagag cgcacatcgc ccacagtccc cgagaagttg gggggagggg tcggcaattg
     7261 aaccggtgcc tagagaaggt ggcgcggggt aaactgggaa agtgatgtcg tgtactggct
     7321 ccgccttttt cccgagggtg ggggagaacc gtatataagt gcagtagtcg ccgtgaacgt
     7381 tctttttcgc aacgggtttg ccgccagaac acaggtaagt gccgtgtgtg gttcccgcgg
     7441 gcctggcctc tttacgggtt atggcccttg cgtgccttga attacttcca cgcccctggc
     7501 tgcagtacgt gattcttgat cccgagcttc gggttggaag tgggtgggag agttcgaggc
     7561 cttgcgctta aggagcccct tcgcctcgtg cttgagttga ggcctggctt gggcgctggg
     7621 gccgccgcgt gcgaatctgg tggcaccttc gcgcctgtct cgctgctttc gataagtctc
     7681 tagccattta aaatttttga tgacctgctg cgacgctttt tttctggcaa gatagtcttg
     7741 taaatgcggg ccaagatctg cacactggta tttcggtttt tggggccgcg ggcggcgacg
     7801 gggcccgtgc gtcccagcgc acatgttcgg cgaggcgggg cctgcgagcg cggccaccga
     7861 gaatcggacg ggggtagtct caagctggcc ggcctgctct ggtgcctggc ctcgcgccgc
     7921 cgtgtatcgc cccgccctgg gcggcaaggc tggcccggtc ggcaccagtt gcgtgagcgg
     7981 aaagatggcc gcttcccggc cctgctgcag ggagctcaaa atggaggacg cggcgctcgg
     8041 gagagcgggc gggtgagtca cccacacaaa ggaaaagggc ctttccgtcc tcagccgtcg
     8101 cttcatgtga ctccacggag taccgggcgc cgtccaggca cctcgattag ttctcgagct
     8161 tttggagtac gtcgtcttta ggttgggggg aggggtttta tgcgatggag tttccccata
     8221 ctgagtgggt ggagactgaa gttaggccag cttggcactt gatgtaattc tccttggaat
     8281 ttgccctttt tgagtttgga tcttggttca ttctcaagcc tcagacagtg gttcaaagtt
     8341 tttttcttcc atttaaggtg tcgtga
//
