LOCUS       sequence-445665-        5972 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cell transfection and expression of M-Ras.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5972)
  AUTHORS   Morstein J, Bowcut V, Fernando M, Yang Y, Zhu L, Jenkins ML, Evans 
            JT, Guiley KZ, Peacock DM, Krahnke S, Lin Z, Taran KA, Huang BJ, 
            Stephen AG, Burke JE, Lightstone FC, Shokat KM
  TITLE     Targeting Ras-, Rho-, and Rab-family GTPases via a conserved cryptic
            pocket.
  JOURNAL   Cell. 2024 Sep 6:S0092-8674(24)00908-5. doi: 
            10.1016/j.cell.2024.08.017.
  PUBMED    39255801
REFERENCE   2  (bases 1 to 5972)
  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: "Cell. 2024 
            Sep 6:S0092-8674(24)00908-5. doi: 10.1016/j.cell.2024.08.017."
COMMENT     Sequence Label: pcDNA3.1(+) M-Ras(WT)
FEATURES             Location/Qualifiers
     source          1..5972
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      917..926
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(1563..1580)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1569..1793
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1839..2267
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1926..1945)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2136..2157
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2276..2296)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2281..2610
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2461..2596
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2523..2542
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2677..3471
                     /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="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2731..2750)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3341..3360
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3645..3766
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3682..3701)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3736..3755
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3815..3831)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3815..3831)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3828..3850)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3839..3855
                     /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(3863..3869,3870..3887,3888..3893))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3908..3929
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4046..4063)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4217..4805)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4297..4316)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4976..5767,5768..5836))
                     /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     5599..5618
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5837..5941)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttgcca ccatggcaac cagcgccgtc cccagtgaca acctccccac
      961 atacaagctg gtggtggtgg gggatggggg tgtgggcaaa agtgccctca ccatccagtt
     1021 tttccagaag atctttgtgc ctgactatga ccccaccatt gaagactcct acctgaaaca
     1081 tacggagatt gacaatcaat gggccatctt ggacgttctg gacacagctg ggcaggagga
     1141 attcagcgcc atgcgggagc aatacatgcg cacgggggat ggcttcctca tcgtctactc
     1201 cgtcactgac aaggccagct ttgagcacgt ggaccgcttc caccagctta tcctgcgcgt
     1261 caaagacagg gagtcattcc cgatgatcct cgtggccaac aaggtcgatt tgatgcactt
     1321 gaggaagatc accagggagc aaggaaaaga aatggcgacc aaacacaata ttccgtacat
     1381 agaaaccagt gccaaggacc cacctctcaa tgtcgacaaa gccttccatg acctcgttag
     1441 agtaattagg caacagattc cggaaaaaag ccagaagaag aagaagaaaa ccaaatggcg
     1501 gggagaccgg gccacaggca cccacaaact gcaatgtgtg atcttgtgaa cccgctgatc
     1561 agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
     1621 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
     1681 gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg
     1741 ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga
     1801 ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt
     1861 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     1921 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     1981 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     2041 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     2101 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     2161 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     2221 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat
     2281 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     2341 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     2401 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     2461 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     2521 tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga
     2581 ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat atccattttc
     2641 ggatctgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac
     2701 gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca
     2761 atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt
     2821 gtcaagaccg acctgtccgg tgccctgaat gaactgcagg acgaggcagc gcggctatcg
     2881 tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga
     2941 agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct
     3001 cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg
     3061 gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg
     3121 gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc
     3181 gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat
     3241 ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac
     3301 tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt
     3361 gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct
     3421 cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc
     3481 tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca
     3541 ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga
     3601 tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag
     3661 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     3721 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac
     3781 cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt
     3841 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg
     3901 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
     3961 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     4021 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     4081 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     4141 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     4201 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     4261 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     4321 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     4381 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     4441 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     4501 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     4561 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     4621 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     4681 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     4741 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     4801 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     4861 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     4921 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     4981 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     5041 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     5101 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     5161 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     5221 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     5281 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     5341 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     5401 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     5461 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     5521 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     5581 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     5641 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     5701 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     5761 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     5821 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     5881 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     5941 gcacatttcc ccgaaaagtg ccacctgacg tc
//
