LOCUS       sequence-227285-        5990 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 5990)
  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 5990)
  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-Rap1b V12
FEATURES             Location/Qualifiers
     source          1..5990
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        125..504
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        505..708
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     659..679
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     753..772
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        753..771
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      803..812
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             812..859
                     /codon_start=1
                     /product="two tandem FLAG(R) epitope tags"
                     /label=2xFLAG
                     /translation="DYKDDDDKDYKDDDDK"
     promoter        complement(1433..1451)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1434..1451)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1471..1488)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1477..1701
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1747..2175
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1834..1853)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2044..2065
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2184..2204)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2189..2518
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2369..2504
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2431..2450
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2585..3379
                     /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(2639..2658)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3249..3268
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3553..3674
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3590..3609)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3644..3663
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3723..3739)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3723..3739)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3736..3758)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3747..3763
                     /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(3771..3777,3778..3795,3796..3801))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3816..3837
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3954..3971)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4125..4713)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4205..4224)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4884..5675,5676..5744))
                     /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     5507..5526
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5745..5849)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5924..5943)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gagtagtgcg cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg
       61 aagaatctgc ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg
      121 cgttgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat
      181 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
      241 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
      301 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
      361 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
      421 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
      481 gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga
      541 tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg
      601 ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg
      661 caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact
      721 agagaaccca ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa
      781 gcttggtacc gagctcggat ccgccaccat ggactacaag gacgacgatg acaaggacta
      841 caaggacgac gatgacaaga attctcgtga gtataagcta gtcgttcttg gctctgtagg
      901 cgttggaaaa tctgctctga ctgtacagtt tgttcaagga atttttgttg aaaaatatga
      961 tcctacgata gaagattctt atagaaagca agttgaagta gatgcacagc agtgtatgct
     1021 tgaaatcttg gatactgcag gaacggaaca attcacagca atgagggatt tgtacatgaa
     1081 aaatggacaa ggctttgcgt tagtttattc catcacagca cagtccacat ttaatgattt
     1141 acaagatctg agagaacaga tacttcgagt taaagacacg gatgatgttc caatgattct
     1201 ggttggtaat aagtgtgact tggaagatga aagagttgta ggaaaggaac aaggtcaaaa
     1261 tctagcaaga cagtggaaca actgtgcatt cttagaatcc tctgcaaaat caaaaataaa
     1321 tgttaatgag atcttttatg acctagtgcg acaaattaac agaaaaactc cagtgcctgg
     1381 gaaggcccgc aaaaagtcat catgtcagct gctttaatct agagggccct attctatagt
     1441 gtcacctaaa tgctagagct cgctgatcag cctcgactgt gccttctagt tgccagccat
     1501 ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc
     1561 tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg
     1621 ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg
     1681 gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctggggc tctagggggt
     1741 atccccacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     1801 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     1861 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     1921 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     1981 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     2041 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
     2101 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     2161 aatttaacgc gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
     2221 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
     2281 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     2341 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     2401 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc
     2461 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
     2521 gctcccggga gcttgtatat ccattttcgg atctgatcaa gagacaggat gaggatcgtt
     2581 tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct
     2641 attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct
     2701 gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga
     2761 actgcaggac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc
     2821 tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg
     2881 gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc
     2941 aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca
     3001 tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga
     3061 cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg cgcgcatgcc
     3121 cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga
     3181 aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca
     3241 ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg
     3301 cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct
     3361 tcttgacgag ttcttctgag cgggactctg gggttcgaaa tgaccgacca agcgacgccc
     3421 aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga
     3481 atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc
     3541 ttcgcccacc ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
     3601 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
     3661 atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt ggcgtaatca
     3721 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     3781 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     3841 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     3901 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     3961 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     4021 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     4081 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     4141 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     4201 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     4261 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     4321 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     4381 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     4441 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     4501 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     4561 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     4621 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     4681 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     4741 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     4801 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     4861 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     4921 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     4981 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     5041 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     5101 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     5161 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     5221 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     5281 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     5341 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     5401 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     5461 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     5521 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     5581 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     5641 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     5701 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     5761 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     5821 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
     5881 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
     5941 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct
//
