LOCUS       sequence-268625-        8259 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of human p115-RhoGEF with N-terminal V5.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8259)
  AUTHORS   de Carcer G, Wachowicz P, Martinez-Martinez S, Oller J, 
            Mendez-Barbero N, Escobar B, Gonzalez-Loyola A, Takaki T, El Bakkali
            A, Camara JA, Jimenez-Borreguero LJ, Bustelo XR, Canamero M, Mulero 
            F, de Los Angeles Sevilla M, Montero MJ, Redondo JM, Malumbres M
  TITLE     Plk1 regulates contraction of postmitotic smooth muscle cells and is
            required for vascular homeostasis.
  JOURNAL   Nat Med. 2017 Aug;23(8):964-974. doi: 10.1038/nm.4364. Epub 2017 Jul
            10.
  PUBMED    28692064
REFERENCE   2  (bases 1 to 8259)
  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"; doi: "10.1038/nm.4364"; 
            journalName: "Nat Med"; date: "2017-08"; volume: "23"; issue: "8"; 
            pages: "964-974"
COMMENT     Sequence Label: pDEST-V5_Hu_p115-RhoGEF(WT)
FEATURES             Location/Qualifiers
     source          1..8259
                     /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             914..955
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             956..976
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     protein_bind    1007..1031
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(3807..3831)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(3850..3867)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3856..4080
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4126..4554
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4213..4232)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4423..4444
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4563..4583)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4568..4897
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4748..4883
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4810..4829
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4964..5758
                     /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(5018..5037)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5628..5647
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5932..6053
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5969..5988)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6023..6042
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6102..6118)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6102..6118)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6115..6137)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6126..6142
                     /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(6150..6156,6157..6174,6175..6180))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6195..6216
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6333..6350)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6504..7092)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6584..6603)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7263..8054,8055..8123))
                     /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     7886..7905
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8124..8228)
                     /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 gctggctagt
      901 taagcttacc atgggtaagc ctatccctaa ccctctcctc ggtctcgatt ctacggaaaa
      961 cctgtatttt cagggcccgg atccgtcgac gaattctgca gatatcacaa gtttgtacaa
     1021 aaaagcaggc tccgcggccg cccccttcac catggaagac ttcgcccgag gggcggcctc
     1081 cccaggcccc tcccggcctg gcctggttcc cgtcagcatc atcggggctg aggatgagga
     1141 ttttgagaac gagctggaga caaactcaga agagcaaaac agccagttcc agagcctgga
     1201 gcaggtgaag cggcgcccag cccacctcat ggccctcctg cagcacgtgg ccctgcagtt
     1261 tgagccagga cccctgcttt gctgtctgca tgccgacatg ctgggctcac tgggccccaa
     1321 ggaggccaag aaggccttcc tggacttcta ccacagcttc ctggagaaga cagcggttct
     1381 ccgggtgccg gtccctccca acgtcgcctt tgaacttgac cgcactaggg ctgacctcat
     1441 ctccgaggat gtccagcggc ggttcgtgca ggaggtggtg caaagccagc aggtagccgt
     1501 gggccggcag ctggaggact tccgttccaa gcggctcatg ggcatgacgc cctgggagca
     1561 ggagctggcc cagctggagg cttgggttgg gcgggaccga gccagctacg aggcccggga
     1621 gcggcacgtg gcggagcggc tgctcatgca cctggaggag atgcaacata ccatctctac
     1681 cgacgaagaa aagagtgctg ccgtggtcaa cgccattggc ctgtacatgc gccaccttgg
     1741 ggtgcggacc aagagtggag acaagaagtc ggggaggaac ttcttccgga aaaaggtgat
     1801 ggggaaccgg cggtcggacg agcctgccaa gaccaagaag gggctgagca gcatcctgga
     1861 tgccgcccgc tggaaccggg gagagcccca ggttccagat tttcgacacc tcaaagcaga
     1921 ggttgatgcc gagaagccag gtgctacaga ccggaaggga ggcgtgggga tgccctctcg
     1981 ggaccggaat atcggggctc ctgggcagga cacccctgga gtctctctgc accctctgtc
     2041 cctggacagc ccagaccggg aaccaggtgc tgacgccccc ctggagctgg gggactcatc
     2101 cccgcagggc ccaatgagcc tggagtcctt ggcgccccca gagagtaccg acgagggggc
     2161 cgaaaccgag agccccgagc ctggagatga gggggagccg gggcggtcgg gactggagct
     2221 tgaaccagaa gagcctcccg gctggcggga actcgtcccc ccagacaccc tgcacagcct
     2281 gcccaagagc caggtgaagc ggcaggaggt catcagcgag ctgctggtga cagaggcggc
     2341 ccacgtgcgc atgctgcggg tgctgcacga cctcttcttc cagcccatgg cagaatgcct
     2401 gttcttcccc ttggaggagc tgcagaacat cttccccagc ctggacgagc tcatcgaggt
     2461 gcattccctg ttcctcgatc gcctgatgaa gcggaggcag gagagtggct acctcatcga
     2521 ggagatcgga gacgtgctgc tggcccggtt tgatggtgct gagggctcct ggttccagaa
     2581 aatctcctcc cgcttctgca gccgccagtc atttgcctta gagcagctca aagccaagca
     2641 acgcaaggac cctcggttct gtgccttcgt gcaggaagct gagagccgcc cgcggtgccg
     2701 ccgcctgcag ctgaaggaca tgatccccac ggagatgcag cggctgacca agtaccccct
     2761 gctcctgcag agcatcgggc agaacacaga agagcccaca gaacgggaga aagtggagct
     2821 ggcagccgag tgctgccggg aaattctaca ccacgtcaac caagccgtgc gtgacatgga
     2881 ggacctgctg aggctcaagg actatcagcg gcgcctggac ttgtcccacc ttcggcagag
     2941 cagcgaccct atgctgagcg agttcaagaa cctggacatc accaagaaga aattggtcca
     3001 cgagggccca ctgacgtggc gggtgactaa ggacaaggca gtggaggtgc atgtgctgct
     3061 gctggacgac ctgctgctgc tgctccagcg ccaggacgag cggctgctgc tcaagtccca
     3121 tagccggaca ctgacgccca cgcccgatgg caagaccatg ctgcggcccg tgctgcggct
     3181 cacctccgcc atgacccgcg aggtggccac cgatcacaaa gccttctacg tcctttttac
     3241 ctgggaccag gaggcccaga tatacgagct ggtggcacag actgtgtcgg agcggaaaaa
     3301 ctggtgtgct ctcatcactg agactgccgg atccctgaaa gtccctgccc ctgcctctcg
     3361 ccctaagccc cggcccagcc cgagcagcac ccgagaaccc ctcctcagca gctctgagaa
     3421 cggcaatggt ggccgagaga cgtctccagc tgatgcccgg accgagagaa tcctcagtga
     3481 cctcctgccc ttctgcagac caggccccga gggccagctc gctgccacgg cccttcggaa
     3541 agtgctgtcc ctgaagcagc ttctgtttcc ggcggaggaa gacaatgggg cggggcctcc
     3601 tcgagatggg gatggggtcc cagggggcgg ccccctgagc ccagcacgga cccaggaaat
     3661 ccaggagaac ctgctcagct tggaggagac catgaagcag ctggaggagt tggaggagga
     3721 attttgccgc ctgagacccc tcctgtctca gcttgggggg aactctgtcc cccagcctgg
     3781 ctgcacttga aagggtgggc gcgccgaccc agctttcttg tacaaagtgg tgataaaccc
     3841 gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg
     3901 tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa
     3961 ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca
     4021 gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg
     4081 cttctgaggc ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg
     4141 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg
     4201 ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc
     4261 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc
     4321 tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga
     4381 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa
     4441 ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg attttgccga
     4501 tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct
     4561 gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     4621 gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc
     4681 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac
     4741 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact
     4801 aatttttttt atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta
     4861 gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc
     4921 cattttcgga tctgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg
     4981 attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca
     5041 acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt
     5101 tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg
     5161 gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga
     5221 agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca
     5281 ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct
     5341 tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac
     5401 tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc
     5461 gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt
     5521 gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt
     5581 catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg
     5641 tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat
     5701 cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc
     5761 gggactctgg ggttcgaaat gaccgaccaa gcgacgccca acctgccatc acgagatttc
     5821 gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc
     5881 tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt
     5941 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca
     6001 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc
     6061 tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg
     6121 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     6181 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     6241 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     6301 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     6361 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     6421 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     6481 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     6541 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     6601 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     6661 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     6721 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     6781 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     6841 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     6901 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     6961 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     7021 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     7081 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     7141 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     7201 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     7261 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     7321 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     7381 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     7441 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     7501 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     7561 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     7621 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     7681 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     7741 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     7801 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     7861 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     7921 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     7981 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     8041 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     8101 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     8161 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     8221 gttccgcgca catttccccg aaaagtgcca cctgacgtc
//
