LOCUS       sequence-227221-        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 N17
FEATURES             Location/Qualifiers
     source          1..5990
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        157..536
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        537..740
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     691..711
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     785..804
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        785..803
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      835..844
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             844..891
                     /codon_start=1
                     /product="two tandem FLAG(R) epitope tags"
                     /label=2xFLAG
                     /translation="DYKDDDDKDYKDDDDK"
     promoter        complement(1465..1483)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1466..1483)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1503..1520)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1509..1733
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1779..2207
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1866..1885)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2076..2097
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2216..2236)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2221..2550
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2401..2536
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2463..2482
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2617..3411
                     /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(2671..2690)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3281..3300
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3585..3706
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3622..3641)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3676..3695
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3755..3771)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3755..3771)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3768..3790)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3779..3795
                     /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(3803..3809,3810..3827,3828..3833))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3848..3869
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3986..4003)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4157..4745)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4237..4256)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4916..5707,5708..5776))
                     /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     5539..5558
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5777..5881)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5956..5975)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta
       61 caacaaggca aggcttgacc gacaattgca tgaagaatct gcttagggtt aggcgttttg
      121 cgctgcttcg cgatgtacgg gccagatata cgcgttgaca ttgattattg actagttatt
      181 aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat
      241 aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa
      301 taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg
      361 agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc
      421 cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct
      481 tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga
      541 tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa
      601 gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc
      661 caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg
      721 aggtctatat aagcagagct ctctggctaa ctagagaacc cactgcttac tggcttatcg
      781 aaattaatac gactcactat agggagaccc aagcttggta ccgagctcgg atccgccacc
      841 atggactaca aggacgacga tgacaaggac tacaaggacg acgatgacaa gaattctcgt
      901 gagtataagc tagtcgttct tggctctgga ggcgttggaa aaaatgctct gactgtacag
      961 tttgttcaag gaatttttgt tgaaaaatat gatcctacga tagaagattc ttatagaaag
     1021 caagttgaag tagatgcaca gcagtgtatg cttgaaatct tggatactgc aggaacggaa
     1081 caattcacag caatgaggga tttgtacatg aaaaatggac aaggctttgc gttagtttat
     1141 tccatcacag cacagtccac atttaatgat ttacaagatc tgagagaaca gatacttcga
     1201 gttaaagaca cggatgatgt tccaatgatt ctggttggta ataagtgtga cttggaagat
     1261 gaaagagttg taggaaagga acaaggtcaa aatctagcaa gacagtggaa caactgtgca
     1321 ttcttagaat cctctgcaaa atcaaaaata aatgttaatg agatctttta tgacctagtg
     1381 cgacaaatta acagaaaaac tccagtgcct gggaaggccc gcaaaaagtc atcatgtcag
     1441 ctgctttaat ctagagggcc ctattctata gtgtcaccta aatgctagag ctcgctgatc
     1501 agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
     1561 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
     1621 gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg
     1681 ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga
     1741 ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt
     1801 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     1861 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     1921 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     1981 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     2041 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     2101 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     2161 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat
     2221 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     2281 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     2341 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     2401 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     2461 tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga
     2521 ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat atccattttc
     2581 ggatctgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac
     2641 gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca
     2701 atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt
     2761 gtcaagaccg acctgtccgg tgccctgaat gaactgcagg acgaggcagc gcggctatcg
     2821 tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga
     2881 agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct
     2941 cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg
     3001 gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg
     3061 gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc
     3121 gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat
     3181 ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac
     3241 tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt
     3301 gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct
     3361 cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc
     3421 tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca
     3481 ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga
     3541 tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag
     3601 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     3661 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac
     3721 cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt
     3781 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg
     3841 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
     3901 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     3961 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     4021 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     4081 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     4141 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     4201 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     4261 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     4321 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     4381 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     4441 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     4501 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     4561 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     4621 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     4681 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     4741 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     4801 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     4861 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     4921 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     4981 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     5041 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     5101 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     5161 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     5221 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     5281 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     5341 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     5401 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     5461 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     5521 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     5581 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     5641 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     5701 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     5761 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     5821 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     5881 gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc gggagatctc ccgatcccct
     5941 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt
//
