LOCUS       sequence-355632-        5802 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Ras Binding Domain split-nTEVP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5802)
  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 5802)
  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-nTEVp
FEATURES             Location/Qualifiers
     source          1..5802
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(585..1376,1377..1445))
                     /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     1208..1227
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1446..1550)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1625..1644)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1811..2190
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2191..2394
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2345..2365
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    2396..2414
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2417..2435
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     2445..2469
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      2561..2570
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3211..3240
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(3296..3313)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3302..3526
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3698..3717)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3849..3896
                     /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             3904..4926
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             3904..4926
                     /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    5056..5177
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5093..5112)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5147..5166
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5226..5242)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5226..5242)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5239..5261)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5250..5266
                     /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(5274..5280,5281..5298,5299..5304))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5319..5340
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5457..5474)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(5628..5802,1..414))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5708..5727)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
       61 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
      121 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
      181 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
      241 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
      301 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
      361 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
      421 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
      481 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
      541 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
      601 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
      661 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
      721 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
      781 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
      841 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
      901 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
      961 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     1021 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     1081 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     1141 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     1201 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     1261 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     1321 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     1381 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     1441 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     1501 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     1561 cgaaaagtgc cacctgacgt cgacggatcg ggagatctcc cgatccccta tggtgcactc
     1621 tcagtacaat ctgctctgat gccgcatagt taagccagta tctgctccct gcttgtgtgt
     1681 tggaggtcgc tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg
     1741 acaattgcat gaagaatctg cttagggtta ggcgttttgc gctgcttcgt acgggccaga
     1801 tatacgcgtt gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta
     1861 gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc
     1921 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
     1981 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
     2041 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
     2101 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
     2161 atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg
     2221 cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg
     2281 agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca
     2341 ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctccct
     2401 atcagtgata gagatctccc tatcagtgat agagatcgtc gacgagctcg tttagtgaac
     2461 cgtcagatcg cctggagacg ccatccacgc tgttttgacc tccatagaag acaccgggac
     2521 cgatccagcc tccggactct agcgtttaaa cttaagcttc gccaccatgg gatccgggga
     2581 gagtttgttt aaggggccaa gggactataa cccaatatca tccactattt gccacctcac
     2641 taacgagagc gatggacata caacctctct ctacgggata ggcttcggtc ctttcatcat
     2701 taccaataag catctgtttc gccgaaacaa cggtactctt ctggttcaat ctcttcatgg
     2761 tgtcttcaag gtgaaaaaca ccactacgct tcagcaacac ctgattgatg gtagggatat
     2821 gataattatc agaatgccga aagatttccc accttttcca cagaagctga aattcaggga
     2881 accgcagaga gaggagagga tttgtttggt aacgaccaac ttccagacgg gctctggcag
     2941 cagcgcggat ccgagtaaga cgtcaaacac aatcagggtt tttctcccca atgaacagcg
     3001 gaccgtggta aacgtccgaa acgggatgag tctccacgac tgtctgatga aagccttgaa
     3061 ggtgaggggg cttcagcctg aatgttgcgc tgtctttcgc cttctccacg aacacaaggg
     3121 cgcaaaggcg aggcttgact ggaatacaga cgctgcaagt ttgattgggg aagaacttca
     3181 ggtcgatttc ctctagctcg aggagggccc gaacaaaaac tcatctcaga agaggatctg
     3241 aatagcgccg tcgaccatca tcataatcat cattgagttt aaacccgctg atcagcctcg
     3301 actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc
     3361 ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt
     3421 ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat
     3481 tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggagat ctatcgatgt
     3541 tctggggatg cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg
     3601 gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     3661 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     3721 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     3781 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
     3841 cgtacctaga agttcctatt ccgaagttcc tattctctag aaagtatagg aacttccttg
     3901 gccaaaaagc ctgaactcac cgcgacgtct gtcgagaagt ttctgatcga aaagttcgac
     3961 agcgtctccg acctgatgca gctctcggag ggcgaagaat ctcgtgcttt cagcttcgat
     4021 gtaggagggc gtggatatgt cctgcgggta aatagctgcg ccgatggttt ctacaaagat
     4081 cgttatgttt atcggcactt tgcatcggcc gcgctcccga ttccggaagt gcttgacatt
     4141 ggggaattca gcgagagcct gacctattgc atctcccgcc gtgcacaggg tgtcacgttg
     4201 caagacctgc ctgaaaccga actgcccgct gttctgcagc cggtcgcgga ggccatggat
     4261 gcgatcgctg cggccgatct tagccagacg agcgggttcg gcccattcgg accgcaagga
     4321 atcggtcaat acactacatg gcgtgatttc atatgcgcga ttgctgatcc ccatgtgtat
     4381 cactggcaaa ctgtgatgga cgacaccgtc agtgcgtccg tcgcgcaggc tctcgatgag
     4441 ctgatgcttt gggccgagga ctgccccgaa gtccggcacc tcgtgcacgc ggatttcggc
     4501 tccaacaatg tcctgacgga caatggccgc ataacagcgg tcattgactg gagcgaggcg
     4561 atgttcgggg attcccaata cgaggtcgcc aacatcttct tctggaggcc gtggttggct
     4621 tgtatggagc agcagacgcg ctacttcgag cggaggcatc cggagcttgc aggatcgccg
     4681 cggctccggg cgtatatgct ccgcattggt cttgaccaac tctatcagag cttggttgac
     4741 ggcaatttcg atgatgcagc ttgggcgcag ggtcgatgcg acgcaatcgt ccgatccgga
     4801 gccgggactg tcgggcgtac acaaatcgcc cgcagaagcg cggccgtctg gaccgatggc
     4861 tgtgtagaag tactcgccga tagtggaaac cgacgcccca gcactcgtcc gagggcaaag
     4921 gaatagcacg tactacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
     4981 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     5041 ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     5101 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     5161 ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa
     5221 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
     5281 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
     5341 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     5401 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
     5461 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     5521 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     5581 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     5641 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     5701 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     5761 accctgccgc ttaccggata cctgtccgcc tttctccctt cg
//
