LOCUS       sequence-445660-        5912 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cell transfection and expression of K-Ras.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5912)
  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 5912)
  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(+) K-Ras(WT)
FEATURES             Location/Qualifiers
     source          1..5912
                     /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"
     CDS             923..1489
                     /codon_start=1
                     /product="human small GTPase proto-oncoprotein"
                     /label=K-Ras
                     /translation="MTEYKLVVVGAGGVGKSALTIQLIQNHFVDEYDPTIEDSYRKQVV
                     IDGETCLLDILDTAGQEEYSAMRDQYMRTGEGFLCVFAINNTKSFEDIHHYREQIKRVK
                     DSEDVPMVLVGNKCDLPSRTVDTKQAQDLARSYGIPFIETSAKTRQGVDDAFYTLVREI
                     RKHKEKMSKDGKKKKKKSKTKCVIM"
     primer_bind     complement(1503..1520)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1509..1733
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1779..2207
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1866..1885)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2076..2097
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2216..2236)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2221..2550
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2401..2536
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2463..2482
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2617..3411
                     /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(2671..2690)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3281..3300
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3585..3706
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3622..3641)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3676..3695
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3755..3771)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3755..3771)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3768..3790)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3779..3795
                     /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(3803..3809,3810..3827,3828..3833))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3848..3869
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3986..4003)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4157..4745)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4237..4256)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4916..5707,5708..5776))
                     /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     5539..5558
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5777..5881)
                     /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 ccatgactga atataaactt gtggtagttg gagctggtgg
      961 cgtaggcaag agtgccttga cgatacagct aattcagaat cattttgtgg acgaatatga
     1021 tccaacaata gaggattcct acaggaagca agtagtaatt gatggagaaa cctgtctctt
     1081 ggatattctc gacacagcag gtcaagagga gtacagtgca atgagggacc agtacatgag
     1141 gactggggag ggctttcttt gtgtatttgc cataaataat actaaatcat ttgaagatat
     1201 tcaccattat agagaacaaa ttaaaagagt taaggactct gaagatgtac ctatggtcct
     1261 agtaggaaat aaatgtgatt tgccttctag aacagtagac acaaaacagg ctcaggactt
     1321 agcaagaagt tatggaattc cttttattga aacatcagca aagacaagac agggtgttga
     1381 tgatgccttc tatacattag ttcgagaaat tcgaaaacat aaagaaaaga tgagcaaaga
     1441 tggtaaaaag aagaaaaaga agtcaaagac aaagtgtgta attatgtaaa cccgctgatc
     1501 agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
     1561 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
     1621 gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg
     1681 ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga
     1741 ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt
     1801 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     1861 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     1921 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     1981 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     2041 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     2101 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     2161 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat
     2221 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     2281 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     2341 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     2401 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     2461 tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga
     2521 ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat atccattttc
     2581 ggatctgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac
     2641 gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca
     2701 atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt
     2761 gtcaagaccg acctgtccgg tgccctgaat gaactgcagg acgaggcagc gcggctatcg
     2821 tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga
     2881 agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct
     2941 cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg
     3001 gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg
     3061 gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc
     3121 gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat
     3181 ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac
     3241 tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt
     3301 gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct
     3361 cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc
     3421 tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca
     3481 ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga
     3541 tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag
     3601 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     3661 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac
     3721 cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt
     3781 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg
     3841 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
     3901 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     3961 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     4021 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     4081 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     4141 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     4201 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     4261 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     4321 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     4381 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     4441 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     4501 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     4561 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     4621 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     4681 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     4741 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     4801 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     4861 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     4921 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     4981 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     5041 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     5101 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     5161 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     5221 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     5281 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     5341 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     5401 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     5461 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     5521 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     5581 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     5641 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     5701 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     5761 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     5821 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     5881 gcacatttcc ccgaaaagtg ccacctgacg tc
//
