LOCUS       sequence-227211-        7427 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of RapGAP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7427)
  AUTHORS   Liu C, Takahashi M, Li Y, Dillon TJ, Kaech S, Stork PJ
  TITLE     The interaction of Epac1 and Ran promotes Rap1 activation at the 
            nuclear envelope.
  JOURNAL   Mol Cell Biol. 2010 Aug;30(16):3956-69. doi: 10.1128/MCB.00242-10. 
            Epub 2010 Jun 14.
  PUBMED    20547757
REFERENCE   2  (bases 1 to 7427)
  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: 
            "10.1128/MCB.00242-10"; journalName: "Mol Cell Biol"; date: 
            "2010-08"; volume: "30"; issue: "16"; pages: "3956-69"
COMMENT     Sequence Label: Flag-RapGAP
FEATURES             Location/Qualifiers
     source          1..7427
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        149..528
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        529..732
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     683..703
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     777..796
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        777..795
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      827..836
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             836..883
                     /codon_start=1
                     /product="two tandem FLAG(R) epitope tags"
                     /label=2xFLAG
                     /translation="DYKDDDDKDYKDDDDK"
     promoter        complement(2894..2912)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2895..2912)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2932..2949)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2938..3162
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3208..3636
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3295..3314)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3505..3526
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3645..3665)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3650..3979
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3830..3965
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3892..3911
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4046..4840
                     /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(4100..4119)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4710..4729
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5014..5135
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5051..5070)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5105..5124
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5184..5200)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5184..5200)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5197..5219)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5208..5224
                     /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(5232..5238,5239..5256,5257..5262))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5277..5298
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5415..5432)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5586..6174)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5666..5685)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6345..7136,7137..7205))
                     /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     6968..6987
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7206..7310)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7385..7404)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca aaatttaagc tacaacaagg
       61 caaggcttga ccgacaattg catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt
      121 cgcgatgtac gggccagata tacgcgttga cattgattat tgactagtta ttaatagtaa
      181 tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg
      241 gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg
      301 tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta
      361 cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt
      421 gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac
      481 tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt gatgcggttt
      541 tggcagtaca tcaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac
      601 cccattgacg tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt
      661 cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat
      721 ataagcagag ctctctggct aactagagaa cccactgctt actggcttat cgaaattaat
      781 acgactcact atagggagac ccaagcttgg taccgagctc ggatccgcca ccatggacta
      841 caaggacgac gatgacaagg actacaagga cgacgatgac aagaattcta ttgagaagat
      901 gcagggaagc aggatggatg aacaacgctg ctccttcccg ccgcccctca aaacagagga
      961 ggactacatt ccatacccga gcgtgcacga ggtcttgggg cgagaaggac ccttccccct
     1021 catcctgctg ccccagtttg ggggctactg gattgagggc accaaccacg aaatcaccag
     1081 catccccgag acagagccac tgcagtcgcc cacaaccaag gtgaagctcg agtgcaaccc
     1141 cacagcccgc atctaccgga agcactttct cggcaaggag catttcaatt actactcact
     1201 ggacgctgcc ctcggccacc ttgtcttctc actcaagtac gatgtcatcg gggaccaaga
     1261 gcacctgcgg ctgctgctca ggaccaagtg ccggacatac catgatgtca tccccatctc
     1321 ctgcctcacc gagttcccta atgttgtcca gatggcaaag ttggtgtgtg aagacgtcaa
     1381 tgtggatcgg ttctatcctg tgctctaccc caaggcttcc cggctcatcg tcacctttga
     1441 cgagcatgtc atcagcaata acttcaagtt tggcgtcatt tatcagaagc ttgggcagac
     1501 ctccgaggaa gaactcttca gcaccaatga ggaaagtccc gctttcgtgg agttccttga
     1561 atttcttggc cagaaggtca aactgcagga ctttaagggg ttccgaggag gcctggacgt
     1621 gacccacggg cagacgggga ccgaatctgt gtactgcaac ttccgcaaca aggagatcat
     1681 gtttcacgtg tccaccaagc tgccatacac ggaaggggac gcccagcagt tgcagcggaa
     1741 gcggcacatc gggaacgaca tcgtggctgt ggtcttccag gatgaaaaca ctcctttcgt
     1801 gcccgacatg atcgcgtcca acttcctgca tgcctacgtc gtggtgcagg ctgagggcgg
     1861 gggccctgat ggccccctct acaaggtctc tgtcactgca agagatgatg tgcccttctt
     1921 tggacccccc ctcccggacc ccgctgtgtt caggaagggg cctgagttcc aggaattttt
     1981 gctgacaaag ctgatcaatg ctgaatatgc ctgctacaag gcagagaagt ttgccaaact
     2041 ggaggagcgg acgcgggccg ccctcctgga gacgctctat gaggaactac acatccacag
     2101 ccagtccatg atgggcttgg gcggcgacga ggacaagatg gagaatggca gtgggggcgg
     2161 cggcttcttt gagtctttca agcgggtcat ccggagccgc agccagtcca tggatgccat
     2221 ggggctgagc aacaagaagc ccaacaccgt gtccaccagc cacagcggga gcttcgcgcc
     2281 caacaacccc gacctggcca aggcggctgg aatatcactg attgtccctg ggaagagccc
     2341 cacgaggaag aagtcgggcc cgttcggctc ccgccgcagc agcgccattg gcatcgagaa
     2401 catacaggag gtgcaggaga agagggagag ccctccggct ggtcagaaga ccccagacag
     2461 cgggcacgtc tcacaggagc ccaagtcgga gaactcatcc actcagagct ccccagagat
     2521 gcccacgacc aagaacagag cggagaccgc agcgcagaga gcagaggcgc tcaaggactt
     2581 ctcccgctcc tcgtccagtg ccagcagctt cgccagcgtg gtggaggaga cggagggtgt
     2641 ggacggagag gacacaggcc tggagagcgt gtcatcctca ggaacacccc acaagcggga
     2701 ctccttcatc tatagcacgt ggctggagga cagtgtcagc accactagtg ggggcagctc
     2761 cccaggcccc tctcgatcac cccacccaga cgccggcaag ttgggggacc ctgcgtgtcc
     2821 cgagatcaag atccagctgg aagcatctga gcagcacatg ccccagctgg gctgttagtc
     2881 tagagggccc tattctatag tgtcacctaa atgctagagc tcgctgatca gcctcgactg
     2941 tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg
     3001 aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga
     3061 gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg
     3121 aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa
     3181 ccagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg
     3241 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     3301 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     3361 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     3421 attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     3481 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
     3541 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     3601 aaaatgagct gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt
     3661 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     3721 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     3781 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct
     3841 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc
     3901 agaggccgag gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg
     3961 aggcctaggc ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca
     4021 agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc
     4081 ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc
     4141 tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga
     4201 cctgtccggt gccctgaatg aactgcagga cgaggcagcg cggctatcgt ggctggccac
     4261 gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct
     4321 gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa
     4381 agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc
     4441 attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct
     4501 tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc
     4561 caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg
     4621 cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct
     4681 gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct
     4741 tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca
     4801 gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgaa
     4861 atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc
     4921 tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc
     4981 ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt
     5041 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
     5101 agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct
     5161 agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     5221 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga
     5281 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     5341 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     5401 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     5461 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     5521 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     5581 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     5641 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     5701 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     5761 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     5821 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     5881 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     5941 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     6001 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     6061 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     6121 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     6181 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     6241 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     6301 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     6361 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     6421 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     6481 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     6541 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     6601 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     6661 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     6721 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     6781 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     6841 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     6901 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     6961 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     7021 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     7081 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     7141 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     7201 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     7261 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     7321 cgaaaagtgc cacctgacgt cgacggatcg ggagatctcc cgatccccta tggtgcactc
     7381 tcagtacaat ctgctctgat gccgcatagt taagccagta tctgctc
//
