LOCUS       sequence-445624-        5283 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein purification of H-Ras (G12C).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5283)
  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 5283)
  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: pProEx H-Ras(G12C) CL
FEATURES             Location/Qualifiers
     source          1..5283
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    230..246
                     /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)."
     primer_bind     235..257
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             279..296
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             318..338
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(973..990)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1143..1229
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1321..1348
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        1367..1458
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1459..1527,1528..2319)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1677..1696)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2490..3078
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2979..2998
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3232..3249
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3326..3754)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3436..3457)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3648..3667
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(3792..3811)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3984..4006
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     promoter        4084..4161
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             4162..5244
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(4181..4200)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 gtttgacagc ttatcatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc
       61 ggaagctgtg gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc
      121 gcactcccgt tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc
      181 tgaaatgagc tgttgacaat taatcatccg gtccgtataa tctgtggaat tgtgagcgga
      241 taacaatttc acacaggaaa cagaccatgt cgtactacca tcaccatcac catcacgatt
      301 acgatatccc aacgaccgaa aacctgtatt ttcagggcgc catgggatcc atgacggaat
      361 ataagctggt ggtggtgggc gcctgcggtg tgggcaagag tgcgctgacc atccagctga
      421 tccagaacca ttttgtggac gaatacgacc ccactataga ggattcctac cggaagcagg
      481 tggtcattga tggggagacg agcctgttgg acatcctgga taccgccggc caggaggagt
      541 acagcgccat gcgggaccag tacatgcgca ccggggaggg cttcctgctg gtgtttgcca
      601 tcaacaacac caagtctttt gaggacatcc accagtacag ggagcagatc aaacgggtga
      661 aggactcgga tgacgtgccc atggtgctgg tggggaacaa gagtgacctg gctgcacgca
      721 ctgtggaatc tcggcaggct caggacctcg cccgaagcta cggcatcccc tacatcgaga
      781 cctcggccaa gacccggcag ggagtggagg atgccttcta cacgttggtg cgtgagatcc
      841 ggcagcactg agaattcaaa ggcctacgtc gacgagctca ctagtcgcgg ccgctttcga
      901 atctagagcc tgcagtctcg aggcatgcgg taccaagctt ggctgttttg gcggatgaga
      961 gaagattttc agcctgatac agattaaatc agaacgcaga agcggtctga taaaacagaa
     1021 tttgcctggc ggcagtagcg cggtggtccc acctgacccc atgccgaact cagaagtgaa
     1081 acgccgtagc gccgatggta gtgtggggtc tccccatgcg agagtaggga actgccaggc
     1141 atcaaataaa acgaaaggct cagtcgaaag actgggcctt tcgttttatc tgttgtttgt
     1201 cggtgaacgc tctcctgagt aggacaaatc cgccgggagc ggatttgaac gttgcgaagc
     1261 aacggcccgg agggtggcgg gcaggacgcc cgccataaac tgccaggcat caaattaagc
     1321 agaaggccat cctgacggat ggcctttttg cgtttctaca aactcttttg tttatttttc
     1381 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
     1441 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt
     1501 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct
     1561 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc
     1621 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta
     1681 tgtggcgcgg tattatcccg tgttgacgcc gggcaagagc aactcggtcg ccgcatacac
     1741 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc
     1801 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac
     1861 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg
     1921 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac
     1981 gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc
     2041 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt
     2101 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga
     2161 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc
     2221 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag
     2281 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca
     2341 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     2401 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     2461 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     2521 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     2581 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt
     2641 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     2701 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     2761 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     2821 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     2881 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     2941 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     3001 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     3061 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     3121 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
     3181 accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca
     3241 gtgagcgagg aagcggaaga gcgcctgatg cggtattttc tccttacgca tctgtgcggt
     3301 atttcacacc gcataatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt
     3361 tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata
     3421 gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac
     3481 gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa
     3541 tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc
     3601 cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg
     3661 aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca
     3721 cccgccgcgc ttaatgcgcc gctacagggc gcgtcccatt cgccattcag gctgctatgg
     3781 tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtacca gtcacgtagc
     3841 gatatcgcta cgtgactggt acactccgct atcgctacgt gactgggtca tggctgcgcc
     3901 ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc
     3961 ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc
     4021 accgaaacgc gcgaggcagc agatcaattc gcgcgcgaag gcgaagcggc atgcatttac
     4081 gttgacacca tcgaatggtg caaaaccttt cgcggtatgg catgatagcg cccggaagag
     4141 agtcaattca gggtggtgaa tgtgaaacca gtaacgttat acgatgtcgc agagtatgcc
     4201 ggtgtctctt atcagaccgt ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa
     4261 acgcgggaaa aagtggaagc ggcgatggcg gagctgaatt acattcccaa ccgcgtggca
     4321 caacaactgg cgggcaaaca gtcgttgctg attggcgttg ccacctccag tctggccctg
     4381 cacgcgccgt cgcaaattgt cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc
     4441 gtggtggtgt cgatggtaga acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat
     4501 cttctcgcgc aacgcgtcag tgggctgatc attaactatc cgctggatga ccaggatgcc
     4561 attgctgtgg aagctgcctg cactaatgtt ccggcgttat ttcttgatgt ctctgaccag
     4621 acacccatca acagtattat tttctcccat gaagacggta cgcgactggg cgtggagcat
     4681 ctggtcgcat tgggtcacca gcaaatcgcg ctgttagcgg gcccattaag ttctgtctcg
     4741 gcgcgtctgc gtctggctgg ctggcataaa tatctcactc gcaatcaaat tcagccgata
     4801 gcggaacggg aaggcgactg gagtgccatg tccggttttc aacaaaccat gcaaatgctg
     4861 aatgagggca tcgttcccac tgcgatgctg gttgccaacg atcagatggc gctgggcgca
     4921 atgcgcgcca ttaccgagtc cgggctgcgc gttggtgcgg atatctcggt agtgggatac
     4981 gacgataccg aagacagctc atgttatatc ccgccgttaa ccaccatcaa acaggatttt
     5041 cgcctgctgg ggcaaaccag cgtggaccgc ttgctgcaac tctctcaggg ccaggcggtg
     5101 aagggcaatc agctgttgcc cgtctcactg gtgaaaagaa aaaccaccct ggcacccaat
     5161 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
     5221 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtaagttagc gcgaattgat
     5281 ctg
//
