LOCUS       sequence-355447-        6391 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  ddRFP sensor of Hras.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6391)
  AUTHORS   Kim J , Lee S, Jung K, Oh WC, Kim N, Son S, Jo Y, Kwon HB,  Heo WD
  TITLE     Intensiometric biosensors visualize the activity of multiple small 
            GTPases in vivo.
  JOURNAL   Nat Commun. 2019 Jan 14;10(1):211. doi: 10.1038/s41467-018-08217-3.
  PUBMED    30643148
REFERENCE   2  (bases 1 to 6391)
  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: "2019-01-14"; volume: "10(1):211. doi"; pages: " 
            10.1038/s41467-018-08217-3"
COMMENT     Sequence Label: pCMV-R-Hras
FEATURES             Location/Qualifiers
     source          1..6391
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             1620..1676
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             2421..2990
                     /codon_start=1
                     /product="human small GTPase proto-oncoprotein"
                     /label=H-Ras
                     /translation="MTEYKLVVVGAGGVGKSALTIQLIQNHFVDEYDPTIEDSYRKQVV
                     IDGETCLLDILDTAGQEEYSAMRDQYMRTGEGFLCVFAINNTKSFEDIHQYREQIKRVK
                     DSDDVPMVLVGNKCDLAARTVESRQAQDLARSYGIPYIETSAKTRQGVEDAFYTLVREI
                     RQHKLRKLNPPDESGPGCMSCKCVLS"
     misc_feature    2999..3055
                     /label=MCS
                     /note="multiple cloning site"
     polyA_signal    3179..3300
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3216..3235)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3270..3289
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3307..3762)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3444..3465)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3656..3675
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3789..3893
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3895..4252
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3895..4086
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3918..3938)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4103..4238
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4287..5081
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4287..5081
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4341..4360)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4951..4970
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5269..5288)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5313..5360
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5689..6277
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6178..6197
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggtaccat ggccaccatc aaagagttca tgcgcttcaa ggtgcgcatg gagggctcca
      661 tgaacggcca cgagttcgag atcgagggcg agggcgaggg ccgcccctac gagggcaccc
      721 agaccgccaa gctgaaggtg accaagggcg gccccctgcc cttcgcctgg gacatcctgt
      781 ccccccagtt catgtacggc tccgaggcgt acgtgaggca ccccgccgac atccccgatt
      841 acaagaagct gcccttcccc gagggcttca agtgggagcg cgtgatgaac ttcgaagacg
      901 gcggtctggt gaccgttacc caggactcct ccctgcagga cggcacgctg atctgcaagg
      961 tgaagatgcg cggcaccaac ttcccccccg acggccccgt aatgcagaag aagaccatgg
     1021 gctgggaggc ctccaccgag atgctgtacc ccgaagacgg cgtgctgaag ggccatagct
     1081 atcaggccct gaagctgaag gacggcggcc actacctggt ggagttcgag accatctaca
     1141 tggccaagaa gcccgtgcaa ctgcccggcg attactgtgt ggacaccaag ctggacatca
     1201 cctcccacaa cgaggactac accatcgtgg aacagtacgg gcgctccgag ggccgccacc
     1261 gtctgggcat ggacgagcgg tacaagttag ggcatggtac aggtagcaca ggcagcggta
     1321 gctccggtac cctgtacaag tccggactca gatctcgagc caccatgcct tctaagacaa
     1381 gcaacactat ccgtgttttc ttgccgaaca agcaaagaac agtggtcaat gtgcgaaatg
     1441 gaatgagctt gcatgactgc cttatgaaag cactcaaggt gaggggcctg caaccagagt
     1501 gctgtgcagt gttcagactt ctccacgaac acaaaggtaa aaaagcacgc ttagattgga
     1561 atactgatgc tgcgtctttg attggagaag aacttcaagt agatttcctg ggatccggcg
     1621 caacaaactt ctctctgctg aaacaagccg gagatgtcga agagaatcct ggaccgatgg
     1681 tgagcaagag cgaggaggtc atcaaagagt tcatgcgctt caaggtgcgc ttggagggct
     1741 ccatgaacgg ccacgagttc gagatcgagg gcgagggcga gggccgcccc tacgagggca
     1801 cccagaccgc caagctgaag gtgaccaagg gcggccccct gcccttcgcc tgggacatcc
     1861 tgtcccccca gatcatgtac ggctccaagg cgtacgtgaa gcaccccgcc gacgtccccg
     1921 attacatgaa gctgtccttc cccgagggct tcaagtggga gcgcgtgatg cacttcgagg
     1981 acggcggtct ggtgaccgtg actcaggaca cctccctgca ggacggcacg ctgatctaca
     2041 aggtgaagat gcgcggcacc aacttccccc ccgacggccc cgtaatgcag aagaagacct
     2101 tgggctggga ttatgccacc gagcgcctgt accccgaaga cggcgtgctg aagggcgagc
     2161 ttctggggcg cctgaagctg aaggacggcg gcctcaacct ggtggagttc aagaccatct
     2221 acatggccaa gaagcccgtg caactgcccg gctactactt cgtggacacc aagctgggca
     2281 tcacctccca caacgaggac tacaccatcg tggaacagta cgagcgctcc gagggccgcc
     2341 accacctggg gcatggcacc ggcagcaccg gcagcggcag ctccggtacc gcctcctccg
     2401 aggacaacaa cctgtacaag atgacggaat ataagctggt ggtggtgggc gccggcggtg
     2461 tgggcaagag tgcgctgacc atccagctga tccagaacca ttttgtggac gaatacgacc
     2521 ccactataga ggattcctac cggaagcagg tggtcattga tggggagacg tgcctgttgg
     2581 acatcctgga taccgccggc caggaggagt acagcgccat gcgggaccag tacatgcgca
     2641 ccggggaggg cttcctgtgt gtgtttgcca tcaacaacac caagtctttt gaggacatcc
     2701 accagtacag ggagcagatc aaacgggtga aggactcgga tgacgtgccc atggtgctgg
     2761 tggggaacaa gtgtgacctg gctgcacgca ctgtggaatc tcggcaggct caggacctcg
     2821 cccgaagcta cggcatcccc tacatcgaga cctcggccaa gacccggcag ggagtggagg
     2881 atgccttcta cacgttggtg cgtgagatcc ggcagcacaa gctgcggaag ctgaaccctc
     2941 ctgatgagag tggccccggc tgcatgagct gcaagtgtgt gctctcctga ccggactcag
     3001 atctcgagct caagcttcga attctgcagt cgacggtacc gcgggcccgg gatccaccgg
     3061 atctagataa ctgatcataa tcagccatac cacatttgta gaggttttac ttgctttaaa
     3121 aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg ttgttgttaa
     3181 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     3241 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     3301 acgcgtaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
     3361 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
     3421 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
     3481 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat
     3541 caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag
     3601 ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga
     3661 agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa
     3721 ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc acttttcggg
     3781 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     3841 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtcctgagg
     3901 cggaaagaac cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc
     3961 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc
     4021 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat
     4081 agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc
     4141 gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga
     4201 gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca aagatcgatc
     4261 aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc
     4321 cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct
     4381 ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg
     4441 acctgtccgg tgccctgaat gaactgcaag acgaggcagc gcggctatcg tggctggcca
     4501 cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc
     4561 tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga
     4621 aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc
     4681 cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc
     4741 ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg
     4801 ccaggctcaa ggcgagcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct
     4861 gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc
     4921 tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc
     4981 ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc
     5041 agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga
     5101 aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt
     5161 ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg
     5221 cggggatctc atgctggagt tcttcgccca ccctaggggg aggctaactg aaacacggaa
     5281 ggagacaata ccggaaggaa cccgcgctat gacggcaata aaaagacaga ataaaacgca
     5341 cggtgttggg tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc actctgtcga
     5401 taccccaccg agaccccatt ggggccaata cgcccgcgtt tcttcctttt ccccacccca
     5461 ccccccaagt tcgggtgaag gcccagggct cgcagccaac gtcggggcgg caggccctgc
     5521 catagcctca ggttactcat atatacttta gattgattta aaacttcatt tttaatttaa
     5581 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     5641 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     5701 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     5761 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     5821 gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     5881 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     5941 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     6001 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     6061 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     6121 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     6181 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     6241 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     6301 acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga
     6361 ttctgtggat aaccgtatta ccgccatgca t
//
