LOCUS       sequence-227762-        5272 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of bovine Rap1b.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5272)
  AUTHORS   Takahashi M, Dillon TJ, Liu C, Kariya Y, Wang Z, Stork PJ
  TITLE     Protein kinase A-dependent phosphorylation of Rap1 regulates its 
            membrane localization and cell migration.
  JOURNAL   J Biol Chem. 2013 Sep 27;288(39):27712-23. doi: 
            10.1074/jbc.M113.466904. Epub 2013 Aug 14.
  PUBMED    23946483
REFERENCE   2  (bases 1 to 5272)
  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: 
            "10.1074/jbc.M113.466904"; journalName: "J Biol Chem"; date: 
            "2013-09-27- 27"; volume: "288"; issue: "39"; pages: "27712-23"
COMMENT     Sequence Label: GFP-Rap1b E63
FEATURES             Location/Qualifiers
     source          1..5272
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        101..404
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        405..608
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     559..579
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      647..656
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(653..655,656..658,659..1369)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(653..655,656..658,659..1369)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(698..719)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(959..978)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1306..1327
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2100..2221
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2137..2156)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2191..2210
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2228..2683)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2365..2386)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2577..2596
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2710..2814
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2816..3173
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2816..3007
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2839..2859)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3024..3159
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3208..4002
                     /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             3208..4002
                     /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(3262..3281)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3872..3891
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4190..4209)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4234..4281
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4610..5198
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5099..5118
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 atcccctgat tctgtggata accgtattac cgccatgcat tagttattaa tagtaatcaa
       61 ttacggggtc attagttcat agcccatata tggagttccg cgttacataa cttacggtaa
      121 atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg
      181 ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt
      241 aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg
      301 tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc
      361 ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc
      421 agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca
      481 ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta
      541 acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa
      601 gcagagctgg tttagtgaac cgtcagatcc gctagcgcta ccggtcgcca ccatggtgag
      661 caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt
      721 aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct
      781 gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac
      841 caccctgacc tacggcgtgc agtgcttcag ccgctacccc gaccacatga agcagcacga
      901 cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga
      961 cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg
     1021 catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga
     1081 gtacaactac aacagccaca acgtctatat catggccgac aagcagaaga acggcatcaa
     1141 ggtgaacttc aagatccgcc acaacatcga ggacggcagc gtgcagctcg ccgaccacta
     1201 ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag
     1261 cacccagtcc gccctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga
     1321 gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagt ccggactcag
     1381 atctcgagct caagcttcga attctcgtga gtataagcta gtcgttcttg gctctggagg
     1441 cgttggaaaa tctgctctga ctgtacagtt tgttcaagga atttttgttg aaaaatatga
     1501 tcctacgata gaagattctt atagaaagca agttgaagta gatgcacagc agtgtatgct
     1561 tgaaatcttg gatactgcag gaacggaaga gttcacagca atgagggatt tgtacatgaa
     1621 aaatggacaa ggctttgcgt tagtttattc catcacagca cagtccacat ttaatgattt
     1681 acaagatctg agagaacaga tacttcgagt taaagacacg gatgatgttc caatgattct
     1741 ggttggtaat aagtgtgact tggaagatga aagagttgta ggaaaggaac aaggtcaaaa
     1801 tctagcaaga cagtggaaca actgtgcatt cttagaatcc tctgcaaaat caaaaataaa
     1861 tgttaatgag atcttttatg acctagtgcg acaaattaac agaaaaactc cagtgcctgg
     1921 gaaggcccgc aaaaagtcat catgtcagct gctttaatct agagggcccg ggatccaccg
     1981 gatctagata actgatcata atcagccata ccacatttgt agaggtttta cttgctttaa
     2041 aaaacctccc acacctcccc ctgaacctga aacataaaat gaatgcaatt gttgttgtta
     2101 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
     2161 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     2221 aacgcgtaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc
     2281 agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag
     2341 accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg
     2401 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca
     2461 tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa
     2521 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg
     2581 aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta
     2641 accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtcaggtgg cacttttcgg
     2701 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     2761 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtcctgag
     2821 gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc
     2881 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt
     2941 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     3001 tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc
     3061 cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tcggcctctg
     3121 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaagatcgat
     3181 caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct
     3241 ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc
     3301 tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc
     3361 gacctgtccg gtgccctgaa tgaactgcaa gacgaggcag cgcggctatc gtggctggcc
     3421 acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg
     3481 ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag
     3541 aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc
     3601 ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt
     3661 cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc
     3721 gccaggctca aggcgagcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc
     3781 tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg
     3841 ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag
     3901 cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg
     3961 cagcgcatcg ccttctatcg ccttcttgac gagttcttct gagcgggact ctggggttcg
     4021 aaatgaccga ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct
     4081 tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc
     4141 gcggggatct catgctggag ttcttcgccc accctagggg gaggctaact gaaacacgga
     4201 aggagacaat accggaagga acccgcgcta tgacggcaat aaaaagacag aataaaacgc
     4261 acggtgttgg gtcgtttgtt cataaacgcg gggttcggtc ccagggctgg cactctgtcg
     4321 ataccccacc gagaccccat tggggccaat acgcccgcgt ttcttccttt tccccacccc
     4381 accccccaag ttcgggtgaa ggcccagggc tcgcagccaa cgtcggggcg gcaggccctg
     4441 ccatagcctc aggttactca tatatacttt agattgattt aaaacttcat ttttaattta
     4501 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     4561 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     4621 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     4681 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     4741 agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     4801 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     4861 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     4921 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     4981 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     5041 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     5101 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     5161 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     5221 tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg tt
//
