LOCUS       sequence-472894-        7128 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Sleeping Beauty transposon plasmid for hydrodynamic tail vein 
            injection (HDTVi) into mice. Ubc promotor driven expression of 
            mVenus and mutant NRAS (NRASG12V)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7128)
  AUTHORS   Chan ASL, Zhu H, Narita M, Cassidy LD, Young ARJ, Bermejo-Rodriguez 
            C, Janowska AT, Chen HC, Gough S, Oshimori N, Zender L, Aitken SJ, 
            Hoare M, Narita M
  TITLE     Titration of RAS alters senescent state and influences tumour 
            initiation.
  JOURNAL   Nature. 2024 Sep;633(8030):678-685. doi: 10.1038/s41586-024-07797-z.
            Epub 2024 Aug 7.
  PUBMED    39112713
REFERENCE   2  (bases 1 to 7128)
  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/s41586-024-07797-z"; journalName: "Nature"; date: 
            "2024-09"; volume: "633"; issue: "8030"; pages: "678-685"
COMMENT     Sequence Label: pT/UBC-mVenus-P2A-NRASG12V
FEATURES             Location/Qualifiers
     source          1..7128
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        474..873
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     1522..1541
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             join(1652..1654,1655..1657,1658..2368)
                     /codon_start=1
                     /product="Venus YFP with monomerizing A206K mutation (Nagai
                     et al., 2002; Kremers et al., 2006)"
                     /label=mVenus
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1697..1718)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1958..1977)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2305..2326
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2378..2434
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     misc_feature    3018..3584
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3178..3195)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3405..3424
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     primer_bind     complement(3834..3851)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3840..4064
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4511..4527)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4511..4527)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4524..4546)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4535..4551
                     /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(4559..4565,4566..4583,4584..4589))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4604..4625
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4742..4759)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4913..5501)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4993..5012)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5672..6463,6464..6532))
                     /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     6295..6314
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6533..6637)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6705..6723
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6761..6783)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6883..6902
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7096..7118
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7110..7127
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7111..7127
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 aattcgagct cggtacccta cagttgaagt cggaagttta catacactta agttggagtc
       61 attaaaactc gtttttcaac tactccacaa atttcttgtt aacaaacaat agttttggca
      121 agtcagttag gacatctact ttgtgcatga cacaagtcat ttttccaaca attgtttaca
      181 gacagattat ttcacttata attcactgta tcacaattcc agtgggtcag aagtttacat
      241 acactaagtt gactgtgcct ttaaacagct tggaaaattc cagaaaatga tgtcatggct
      301 ttagaagctt ctgatagact aattgacatc atttgagtca attggaggtg tacctgtgga
      361 tgtatttcaa ggcgatgtac gggccagata tacgcgttga cattgattat tgactagtaa
      421 cggccgccag tgtgctggaa ttctgcagat atcaagctta tcgatggtgc agcggcctcc
      481 gcgccgggtt ttggcgcctc ccgcgggcgc ccccctcctc acggcgagcg ctgccacgtc
      541 agacgaaggg cgcagcgagc gtcctgatcc ttccgcccgg acgctcagga cagcggcccg
      601 ctgctcataa gactcggcct tagaacccca gtatcagcag aaggacattt taggacggga
      661 cttgggtgac tctagggcac tggttttctt tccagagagc ggaacaggcg aggaaaagta
      721 gtcccttctc ggcgattctg cggagggatc tccgtggggc ggtgaacgcc gatgattata
      781 taaggacgcg ccgggtgtgg cacagctagt tccgtcgcag ccgggatttg ggtcgcggtt
      841 cttgtttgtg gatcgctgtg atcgtcactt ggtgagtagc gggctgctgg gctggccggg
      901 gctttcgtgg ccgccgggcc gctcggtggg acggaagcgt gtggagagac cgccaagggc
      961 tgtagtctgg gtccgcgagc aaggttgccc tgaactgggg gttgggggga gcgcagcaaa
     1021 atggcggctg ttcccgagtc ttgaatggaa gacgcttgtg aggcgggctg tgaggtcgtt
     1081 gaaacaaggt ggggggcatg gtgggcggca agaacccaag gtcttgaggc cttcgctaat
     1141 gcgggaaagc tcttattcgg gtgagatggg ctggggcacc atctggggac cctgacgtga
     1201 agtttgtcac tgactggaga actcggtttg tcgtctgttg cgggggcggc agttatggcg
     1261 gtgccgttgg gcagtgcacc cgtacctttg ggagcgcgcg ccctcgtcgt gtcgtgacgt
     1321 cacccgttct gttggcttat aatgcagggt ggggccacct gccggtaggt gtgcggtagg
     1381 cttttctccg tcgcaggacg cagggttcgg gcctagggta ggctctcctg aatcgacagg
     1441 cgccggacct ctggtgaggg gagggataag tgaggcgtca gtttctttgg tcggttttat
     1501 gtacctatct tcttaagtag ctgaagctcc ggttttgaac tatgcgctcg gggttggcga
     1561 gtgtgttttg tgaagttttt taggcacctt ttgaaatgta atcatttggg tcaatatgta
     1621 attttcagtg ttagactagt aaagcggccg catggtgagc aagggcgagg agctgttcac
     1681 cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt
     1741 gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagc tgatctgcac
     1801 caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgggct acggcctgca
     1861 gtgcttcgcc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc
     1921 cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg
     1981 cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga
     2041 cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa
     2101 cgtctatatc accgccgaca agcagaagaa cggcatcaag gccaacttca agatccgcca
     2161 caacatcgag gacggcggcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg
     2221 cgacggcccc gtgctgctgc ccgacaacca ctacctgagc taccagtcca agctgagcaa
     2281 agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat
     2341 cactctcggc atggacgagc tgtacaaagg aagcggagct actaacttca gcctgctgaa
     2401 gcaggctgga gacgtggagg agaaccctgg acctatgact gagtacaaac tggtggtggt
     2461 tggagcagtt ggtgttggga aaagcgcact gacaatccag ctaatccaga accactttgt
     2521 agatgaatat gatcccacca tagaggattc ttacagaaaa caagtggtta tagatggtga
     2581 aacctgtttg ttggacatac tggatacagc tggacaagaa gagtacagtg ccatgagaga
     2641 ccaatacatg aggacaggcg aaggcttcct ctgtgtattt gccatcaata atagcaagtc
     2701 atttgcggat attaacctct acagggaaca gattaagcga gtaaaagact cggatgatgt
     2761 acctatggtg ctagtgggaa acaagtgtga tttgccaaca aggacagttg atacaaaaca
     2821 agcccacgaa ctggccaaga gttacgggat tccattcatt gaaacctcag ccaagaccag
     2881 acagggtgtt gaagatgctt tttacacact ggtaagagaa atacgccagt accgaatgaa
     2941 aaaactcaac agcagtgatg atgggactca gggttgtatg ggattgccat gtgtggtgat
     3001 gtaagcggcc aacgaattcc gccccccccc ctaacgttac tggccgaagc cgcttggaat
     3061 aaggccggtg tgcgtttgtc tatatgttat tttccaccat attgccgtct tttggcaatg
     3121 tgagggcccg gaaacctggc cctgtcttct tgacgagcat tcctaggggt ctttcccctc
     3181 tcgccaaagg aatgcaaggt ctgttgaatg tcgtgaagga agcagttcct ctggaagctt
     3241 cttgaagaca aacaacgtct gtagcgaccc tttgcaggca gcggaacccc ccacctggcg
     3301 acaggtgcct ctgcggccaa aagccacgtg tataagatac acctgcaaag gcggcacaac
     3361 cccagtgcca cgttgtgagt tggatagttg tggaaagagt caaatggctc tcctcaagcg
     3421 tattcaacaa ggggctgaag gatgcccaga aggtacccca ttgtatggga tctgatctgg
     3481 ggcctcggtg cacatgcttt acatgtgttt agtcgaggtt aaaaaaacgt ctaggccccc
     3541 cgaaccacgg ggacgtggtt ttcctttgaa aaacacgatg ataagcttga tcacgcgtct
     3601 cgagatatca gtggtccagg ctctagtttt gactcaacaa tatcaccagc tgaagcctat
     3661 agagtacgag ccatagataa aataaaagat tttatttagt ctccagaaaa aggggggaat
     3721 gaaagacccc acctgtaggt ttggcaagct agcattcccg ggaaaactca acagcagtga
     3781 tgatgggact cagggttgta tgggattgcc atgtgtggtg atgtaaggat cagcctcgac
     3841 tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct
     3901 ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct
     3961 gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg
     4021 ggaagacaat agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag
     4081 aaccagccat cacaaagctc tgacctcaat cctatagaaa ggaggaatga gccaaaattc
     4141 acccaactta ttgtgggaag cttgtggaag gctactcgaa atgtttgacc caagttaaac
     4201 aatttaaagg caatgctacc aaatactaat tgagtgtatg ttaacttctg acccactggg
     4261 aatgtgatga aagaaataaa agctgaaatg aatcattctc tctactatta ttctgatatt
     4321 tcacattctt aaaataaagt ggtgatccta actgacctta agacagggaa tctttactcg
     4381 gattaaatgt caggaattgt gaaaaagtga gtttaaatgt atttggctaa ggtgtatgta
     4441 aacttccgac ttcaactgta ggggatcctc tagagtcgac ctgcaggcat gcaagcttgg
     4501 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
     4561 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
     4621 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     4681 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     4741 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     4801 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     4861 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     4921 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     4981 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     5041 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     5101 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     5161 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     5221 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     5281 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     5341 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     5401 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     5461 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     5521 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     5581 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     5641 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     5701 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     5761 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     5821 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     5881 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     5941 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     6001 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     6061 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     6121 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     6181 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     6241 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     6301 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     6361 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     6421 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     6481 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     6541 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     6601 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     6661 ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga
     6721 ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc
     6781 cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg
     6841 cgtcagcggg tgttggcggg tgtcggggct ggcttaacta tgcggcatca gagcagattg
     6901 tactgagagt gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc
     6961 gcatcaggcg ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg
     7021 cctcttcgct attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg
     7081 taacgccagg gttttcccag tcacgacgtt gtaaaacgac ggccagtg
//
