LOCUS       Exported                7348 bp ds-DNA     circular SYN 17-JAN-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7348)
  AUTHORS   MIT
  TITLE     Direct Submission
  JOURNAL   Exported Tuesday, Jan 21, 2025 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7348
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     109..128
                     /label=oSEQ054 pMXs-fwd-seq
     primer_bind     complement(198..211)
                     /label=oSEQ013 flipped_WPRE_rev
     protein_bind    222..246
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(223..241)
                     /label=?oSEQ049 attR1-rev
     misc_feature    289..306
                     /label=MCS
     CDS             319..864
                     /codon_start=1
                     /product="human O6-alkylguanine-DNA-alkyltransferase 
                     variant that forms covalent bonds with benzylguanine 
                     derivatives"
                     /label=SNAP-tag(R)
                     /note="fast-reacting variant, optimized for mammalian 
                     expression"
                     /translation="MDKDCEMKRTTLDSPLGKLELSGCEQGLHRIIFLGKGTSAADAVE
                     VPAPAAVLGGPEPLMQATAWLNAYFHQPEAIEEFPVPALHHPVFQQESFTRQVLWKLLK
                     VVKFGEVISYSHLAALAGNPAATAAVKTALSGNPVPILIPCHRVVQGDLDVGGYEGGLA
                     VKEWLLAHEGHRLGKPGLG"
     CDS             880..1188
                     /codon_start=1
                     /label=p53DD
                     /translation="MTAMEESQSDISLKRALPTCTSASPPQKKKPLDGEYFTLKIRGRK
                     RFEMFRELNEALELKDAHATEESGDSRAHSSYLKTKKGQSTSRHKKTMVKKVGPDSD"
     misc_feature    1202..1740
                     /label=IRES
     primer_bind     complement(1345..1362)
                     /label=oSEQ040 IRES_rev
     primer_bind     1572..1591
                     /label=oSEQ039 IRES_fwd
     CDS             1746..2315
                     /codon_start=1
                     /product="human oncoprotein generated by the G12V mutation 
                     in the small GTPase H-Ras"
                     /label=H-Ras (G12V)
                     /translation="MTEYKLVVVGAVGVGKSALTIQLIQNHFVDEYDPTIEDSYRKQVV
                     IDGETCLLDILDTAGQEEYSAMRDQYMRTGEGFLCVFAINNTKSFEDIHQYREQIKRVK
                     DSDDVPMVLVGNKCDLAARTVESRQAQDLARSYGIPYIETSAKTRQGVEDAFYTLVREI
                     RQHKLRKLNPPDESGPGCMSCKCVLS"
     primer_bind     2201..2219
                     /label=oSEQ092 HRas-mid_fwd
     primer_bind     complement(2294..2339)
                     /label=P2A-hRAS-Rev
                     /note="Melting Temp - 66 C"
     primer_bind     complement(2297..2339)
                     /label=P2A-hRAS-Rev-v02
     protein_bind    complement(2329..2353)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    2370..2958
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2536..2555)
                     /label=oSEQ034 WPRE_rev
     primer_bind     2793..2813
                     /label=oSEQ048 phage-dest-fwd-2
     CDS             complement(2841..2852)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     2955..2972
                     /label=oSEQ013 flipped_WPRE_rev
     primer_bind     complement(2997..3015)
                     /label=oSEQ055 pMXs-rev-seq
     LTR             3138..3607
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     rep_origin      complement(3908..4496)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3988..4007)
                     /label=oSEQ067 pBR322-ori_fwd
     CDS             complement(4660..5520)
                     /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     5283..5302
                     /label=oSEQ066 Amp_rev
     primer_bind     complement(5496..5520)
                     /label=oSEQ083 Amp-fwd
     promoter        complement(5521..5625)
                     /gene="bla"
                     /label=AmpR promoter
     LTR             5655..6248
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     misc_feature    6311..6668
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             6727..7143
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     complement(6921..6940)
                     /label=oSEQ082 pMX-gag-rev
     misc_feature    join(7153..7348,1..179)
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
ORIGIN
        1 tcggcgacac agtgtgggtc cgccgacacc agactaagaa cctagaacct cgctggaaag
       61 gaccttacac agtcctgctg accaccccca ccgccctcaa agtagacggc atcgcagctt
      121 ggatacacgc cgcccacgtg aaggctgccg accccggggg tggaccatcc tctagactgc
      181 cggatctagc tagttaatcg acggtatcga taagctatca aacaagtttg tacaaaaaag
      241 caggctccac catgggaacc aattcagtcg actggatccg gtactactga attcaagctt
      301 ggatcccaat gtgccaccat ggacaaagac tgcgaaatga agcgcaccac cctggatagc
      361 cctctgggca agctggaact gtctgggtgc gaacagggcc tgcaccgtat catcttcctg
      421 ggcaaaggaa catctgccgc cgacgccgtg gaagtgcctg ccccagccgc cgtgctgggc
      481 ggaccagagc cactgatgca ggccaccgcc tggctcaacg cctactttca ccagcctgag
      541 gccatcgagg agttccctgt gccagccctg caccacccag tgttccagca ggagagcttt
      601 acccgccagg tgctgtggaa actgctgaaa gtggtgaagt tcggagaggt catcagctac
      661 agccacctgg ccgccctggc cggcaatccc gccgccaccg ccgccgtgaa aaccgccctg
      721 agcggaaatc ccgtgcccat tctgatcccc tgccaccggg tggtgcaggg cgacctggac
      781 gtggggggct acgagggcgg gctcgccgtg aaagagtggc tgctggccca cgagggccac
      841 agactgggca agcctgggct gggtcctgca ggcggatcca tgactgccat ggaggagtca
      901 cagtcggata tcagcctcaa gagagcgctg cccacctgca caagcgcctc tcccccgcaa
      961 aagaaaaaac cacttgatgg agagtatttc accctcaaga tccgcgggcg taaacgcttc
     1021 gagatgttcc gggagctgaa tgaggcctta gagttaaagg atgcccatgc tacagaggag
     1081 tctggagaca gcagggctca ctccagctac ctgaagacca agaagggcca gtctacttcc
     1141 cgccataaaa aaacaatggt caagaaagtg gggcctgact cagactgaca acaccatgga
     1201 gcgttactgg ccgaagccgc ttggaataag gccggtgtgc gtttgtctat atgttatttt
     1261 ccaccatatt gccgtctttt ggcaatgtga gggcccggaa acctggccct gtcttcttga
     1321 cgagcattcc taggggtctt tcccctctcg ccaaaggaat gcaaggtctg ttgaatgtcg
     1381 tgaaggaagc agttcctctg gaagcttctt gaagacaaac aacgtctgta gcgacccttt
     1441 gcaggcagcg gaacccccca cctggcgaca ggtgcctctg cggccaaaag ccacgtgtat
     1501 aagatacacc tgcaaaggcg gcacaacccc agtgccacgt tgtgagttgg atagttgtgg
     1561 aaagagtcaa atggctctcc tcaagcgtat tcaacaaggg gctgaaggat gcccagaagg
     1621 taccccattg tatgggatct gatctggggc ctcggtgcac atgctttaca tgtgtttagt
     1681 cgaggttaaa aaaacgtcta ggccccccga accacgggga cgtggttttc ctttgaaaaa
     1741 caggaatgac cgaatacaag ctggtggtgg tgggcgccgt cggtgtgggc aagagtgcgc
     1801 tgaccatcca gctgatccag aaccattttg tggacgaata cgaccccact atagaggatt
     1861 cctaccggaa gcaggtggtc attgatgggg agacgtgcct gttggacatc ctggataccg
     1921 ccggccagga ggagtacagc gccatgcggg accagtacat gcgcaccggg gagggcttcc
     1981 tgtgtgtgtt tgccatcaac aacaccaagt cttttgagga catccaccag tacagggagc
     2041 agatcaaacg ggtgaaggac tcggatgacg tgcccatggt gctggtgggg aacaagtgtg
     2101 acctggctgc acgcactgtg gaatctcggc aggctcagga cctcgcccga agctacggca
     2161 tcccctacat cgaaacctcg gccaagaccc ggcagggagt ggaggatgcc ttctacacgt
     2221 tggtgcgtga gatccggcag cacaagctgc ggaagctgaa ccctcctgat gagagtggcc
     2281 ccggctgcat gagctgcaag tgtgtgctct cctaacgctg atatctagac ccagctttct
     2341 tgtacaaagt ggttcgatag cttatcgata atcaacctct ggattacaaa atttgtgaaa
     2401 gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa
     2461 tgcctttgta tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat
     2521 cctggttgct gtctctttat gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt
     2581 gcactgtgtt tgctgacgca acccccactg gttggggcat tgccaccacc tgtcagctcc
     2641 tttccgggac tttcgctttc cccctcccta ttgccacggc ggaactcatc gccgcctgcc
     2701 ttgcccgctg ctggacaggg gctcggctgt tgggcactga caattccgtg gtgttgtcgg
     2761 ggaaatcatc gtcctttcct tggctgctcg cctgtgttgc cacctggatt ctgcgcggga
     2821 cgtccttctg ctacgtccct tcggccctca atccagcgga ccttccttcc cgcggcctgc
     2881 tgccggctct gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt cggatctccc
     2941 tttgggccgc ctccccgcat cgataccgtc gacgataaaa taaaagattt tatttagtct
     3001 ccagaaaaag gggggaatga aagaccccac ctgtaggttt ggcaagctag cttaagtaac
     3061 gccattttgc aaggcatgga aaaatacata actgagaata gagaagttca gatcaaggtc
     3121 aggaacagat ggaacagctg aatatgggcc aaacaggata tctgtggtaa gcagttcctg
     3181 ccccggctca gggccaagaa cagatggaac agctgaatat gggccaaaca ggatatctgt
     3241 ggtaagcagt tcctgccccg gctcagggcc aagaacagat ggtccccaga tgcggtccag
     3301 ccctcagcag tttctagaga accatcagat gtttccaggg tgccccaagg acctgaaatg
     3361 accctgtgcc ttatttgaac taaccaatca gttcgcttct cgcttctgtt cgcgcgcttc
     3421 tgctccccga gctcaataaa agagcccaca acccctcact cggggcgcca gtcctccgat
     3481 tgactgagtc gcccgggtac ccgtgtatcc aataaaccct cttgcagttg catccgactt
     3541 gtggtctcgc tgttccttgg gagggtctcc tctgagtgat tgactacccg tcagcggggg
     3601 tctttcacat gcagcatgta tcaaaattaa tttggttttt tttcttaagt atttacatta
     3661 aatggccata gttgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     3721 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     3781 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     3841 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     3901 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     3961 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     4021 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     4081 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     4141 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     4201 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     4261 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     4321 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     4381 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     4441 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     4501 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     4561 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttgcgg ccggccgcaa
     4621 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     4681 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     4741 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     4801 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     4861 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     4921 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     4981 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     5041 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     5101 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     5161 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     5221 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     5281 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     5341 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     5401 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     5461 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     5521 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     5581 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     5641 agtgccacct gcataatgaa agaccccacc tgtaggtttg gcaagctagc ttaagtaacg
     5701 ccattttgca aggcatggaa aaatacataa ctgagaatag aaaagttcag atcaaggtca
     5761 ggaacagatg gaacagctga atatgggcca aacaggatat ctgtggtaag cagttcctgc
     5821 cccggctcag ggccaagaac agatggaaca gctgaatatg ggccaaacag gatatctgtg
     5881 gtaagcagtt cctgccccgg ctcagggcca agaacagatg gtccccagat gcggtccagc
     5941 cctcagcagt ttctagagaa ccatcagatg tttccagggt gccccaagga cctgaaatga
     6001 ccctgtgcct tatttgaact aaccaatcag ttcgcttctc gcttctgttc gcgcgcttct
     6061 gctccccgag ctcaataaaa gagcccacaa cccctcactc ggcgcgccag tcctccgatt
     6121 gactgagtcg cccgggtacc cgtgtatcca ataaaccctc ttgcagttgc atccgacttg
     6181 tggtctcgct gttccttggg agggtctcct ctgagtgatt gactacccgt cagcgggggt
     6241 ctttcatttg ggggctcgtc cgggatcggg agacccctgc ccagggacca ccgacccacc
     6301 accgggaggt aagctggcca gcaacttatc tgtgtctgtc cgattgtcta gtgtctatga
     6361 ctgattttat gcgcctgcgt cggtactagt tagctaacta gctctgtatc tggcggaccc
     6421 gtggtggaac tgacgagttc ggaacacccg gccgcaaccc tgggagacgt cccagggact
     6481 tcgggggccg tttttgtggc ccgacctgag tccaaaaatc ccgatcgttt tggactcttt
     6541 ggtgcacccc cctaatagga gggatatgtg gttctggtag gagacgagaa cctaaaacag
     6601 ttcccgcctc cgtctgaatt tttgctttcg gtttgggacc gaagccgcgc cgcgcgtctt
     6661 gtctgctgca gcatcgttct gtgttgtctc tgtctgactg tgtttctgta tttgtctgaa
     6721 aattagggcc agactgttac cactccctta agtttgacct taggtcactg gaaagatgtc
     6781 gagcggatcg ctcacaacca gtcggtagat gtcaagaaga gacgttgggt taccttctgc
     6841 tctgcagaat ggccaacctt taacgtcgga tggccgcgag acggcacctt taaccgagac
     6901 ctcatcaccc aggttaagat caaggtcttt tcacctggcc cgcatggaca cccagaccag
     6961 gtcccctaca tcgtgacctg ggaagccttg gcttttgacc cccctccctg ggtcaagccc
     7021 tttgtacacc ctaagcctcc gcctcctctt cctccatccg ccccgtctct cccccttgaa
     7081 cctcctcgtt cgaccccgcc tcgatcctcc ctttatccag ccctcactcc ttctctaggc
     7141 gcccccatat ggccatatga gatcttatat ggggcacccc cgccccttgt aaacttccct
     7201 gaccctgaca tgacaagagt tactaacagc ccctctctcc aagctcactt acaggctctc
     7261 tacttagtcc agcacgaagt ctggagacct ctggcggcag cctaccaaga acaactggac
     7321 cgaccggtgg tacctcaccc ttaccgag
//
