LOCUS       sequence-356811-        5799 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mutant Ras Binding Domain split-cTEVP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5799)
  AUTHORS   Vlahos AE, Kang J, Aldrete CA, Zhu R, Chong LS, Elowitz MB, Gao XJ
  TITLE     Protease-controlled secretion and display of intercellular signals.
  JOURNAL   Nat Commun. 2022 Feb 17;13(1):912. doi: 10.1038/s41467-022-28623-y.
  PUBMED    35177637
REFERENCE   2  (bases 1 to 5799)
  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: "Nat 
            Commun."; date: "2022-02-17"; volume: "13(1):912. doi"; pages: " 
            10.1038/s41467-022-28623-y"
COMMENT     Sequence Label: CMVTO-RBDK65E-R89L-cTEVp
FEATURES             Location/Qualifiers
     source          1..5799
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        116..495
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        496..699
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     650..670
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    701..719
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    722..740
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     750..774
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      866..875
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1513..1542
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(1598..1615)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1604..1828
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2000..2019)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2151..2198
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             2206..3228
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             2206..3228
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    3358..3479
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3395..3414)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3449..3468
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3528..3544)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3528..3544)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3541..3563)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3552..3568
                     /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(3576..3582,3583..3600,3601..3606))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3621..3642
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3759..3776)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3930..4518)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4010..4029)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4689..5480,5481..5549))
                     /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     5312..5331
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5550..5654)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5729..5748)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt gaccgacaat tgcatgaaga
       61 atctgcttag ggttaggcgt tttgcgctgc ttcgtacggg ccagatatac gcgttgacat
      121 tgattattga ctagttatta atagtaatca attacggggt cattagttca tagcccatat
      181 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac
      241 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc
      301 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
      361 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
      421 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
      481 atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt
      541 gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac
      601 caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc
      661 ggtaggcgtg tacggtggga ggtctatata agcagagctc tccctatcag tgatagagat
      721 ctccctatca gtgatagaga tcgtcgacga gctcgtttag tgaaccgtca gatcgcctgg
      781 agacgccatc cacgctgttt tgacctccat agaagacacc gggaccgatc cagcctccgg
      841 actctagcgt ttaaacttaa gcttcgccac catggatccg agtaagacgt caaacacaat
      901 cagggttttt ctccccaatg aacagcggac cgtggtaaac gtccgaaacg ggatgagtct
      961 ccacgactgt ctgatgaaag ccttgaaggt gctggggctt cagcctgaat gttgcgctgt
     1021 ctttcgcctt ctccacgaac acaagggcgc aaaggcgagg cttgactgga atacagacgc
     1081 tgcaagtttg attggggaag aacttcaggt cgatttcctc accggtggct cagggagcaa
     1141 gagtatgagt tctatggtgt ccgacactag ctgcacgttc ccctcaagtg atgggatatt
     1201 ctggaaacac tggatacaga ctaaagacgg acagtgtgga agcccattgg tttccacccg
     1261 agatggtttt atcgtgggta tccatagcgc ctctaatttc acaaacacga acaactattt
     1321 cacttcagtg cccaaaaact ttatggagct gctcacaaac caagaagcgc agcagtgggt
     1381 aagcggttgg agacttaacg ctgactcagt tctctggggg gggcacaaag tattcatggt
     1441 aaagccagag gagccattcc aaccagtcaa agaagccaca caacttatga acagctagct
     1501 cgaggagggc ccgaacaaaa actcatctca gaagaggatc tgaatagcgc cgtcgaccat
     1561 catcataatc atcattgagt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc
     1621 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     1681 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
     1741 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
     1801 catgctgggg atgcggtggg ctctatggag atctatcgat gttctgggga tgcggtgggc
     1861 tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc
     1921 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     1981 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     2041 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     2101 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtaccta gaagttccta
     2161 ttccgaagtt cctattctct agaaagtata ggaacttcct tggccaaaaa gcctgaactc
     2221 accgcgacgt ctgtcgagaa gtttctgatc gaaaagttcg acagcgtctc cgacctgatg
     2281 cagctctcgg agggcgaaga atctcgtgct ttcagcttcg atgtaggagg gcgtggatat
     2341 gtcctgcggg taaatagctg cgccgatggt ttctacaaag atcgttatgt ttatcggcac
     2401 tttgcatcgg ccgcgctccc gattccggaa gtgcttgaca ttggggaatt cagcgagagc
     2461 ctgacctatt gcatctcccg ccgtgcacag ggtgtcacgt tgcaagacct gcctgaaacc
     2521 gaactgcccg ctgttctgca gccggtcgcg gaggccatgg atgcgatcgc tgcggccgat
     2581 cttagccaga cgagcgggtt cggcccattc ggaccgcaag gaatcggtca atacactaca
     2641 tggcgtgatt tcatatgcgc gattgctgat ccccatgtgt atcactggca aactgtgatg
     2701 gacgacaccg tcagtgcgtc cgtcgcgcag gctctcgatg agctgatgct ttgggccgag
     2761 gactgccccg aagtccggca cctcgtgcac gcggatttcg gctccaacaa tgtcctgacg
     2821 gacaatggcc gcataacagc ggtcattgac tggagcgagg cgatgttcgg ggattcccaa
     2881 tacgaggtcg ccaacatctt cttctggagg ccgtggttgg cttgtatgga gcagcagacg
     2941 cgctacttcg agcggaggca tccggagctt gcaggatcgc cgcggctccg ggcgtatatg
     3001 ctccgcattg gtcttgacca actctatcag agcttggttg acggcaattt cgatgatgca
     3061 gcttgggcgc agggtcgatg cgacgcaatc gtccgatccg gagccgggac tgtcgggcgt
     3121 acacaaatcg cccgcagaag cgcggccgtc tggaccgatg gctgtgtaga agtactcgcc
     3181 gatagtggaa accgacgccc cagcactcgt ccgagggcaa aggaatagca cgtactacga
     3241 gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac
     3301 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac
     3361 ttgtttattg cagcttataa tagttacaaa taaagcaata gcatcacaaa tttcacaaat
     3421 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     3481 catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt
     3541 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     3601 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     3661 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     3721 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     3781 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     3841 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     3901 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     3961 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     4021 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     4081 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     4141 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     4201 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     4261 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     4321 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     4381 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     4441 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     4501 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     4561 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     4621 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     4681 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     4741 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     4801 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     4861 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     4921 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     4981 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     5041 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     5101 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     5161 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     5221 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     5281 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     5341 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     5401 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     5461 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     5521 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     5581 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     5641 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtcgacgg atcgggagat
     5701 ctcccgatcc cctatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc
     5761 agtatctgct ccctgcttgt gtgttggagg tcgctgagt
//
