LOCUS       sequence-166557-        6661 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  acute activation of Rho through rapamycin-induced translocation to 
            membrane-localized FRB.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6661)
  AUTHORS   Hayer A, Shao L, Chung M, Joubert LM, Yang HW, Tsai FC, Bisaria A, 
            Betzig E, Meyer T
  TITLE     Engulfed cadherin fingers are polarized junctional structures 
            between collectively migrating endothelial cells.
  JOURNAL   Nat Cell Biol. 2016 Dec;18(12):1311-1323. doi: 10.1038/ncb3438. Epub
            2016 Nov 14.
  PUBMED    27842057
REFERENCE   2  (bases 1 to 6661)
  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/ncb3438"; 
            journalName: "Nat Cell Biol"; date: "2016-12"; volume: "18"; issue: 
            "12"; pages: "1311-1323"
COMMENT     Sequence Label: pCAG-mCherry-FKBP-GEF(ARHGEF1)
FEATURES             Location/Qualifiers
     source          1..6661
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        386..661
                     /label=chicken beta-actin promoter
     intron          662..1670
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1594..1616
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1678..1697
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             1725..2432
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1870..1888)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2138..2157
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             2445..2765
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     primer_bind     complement(3685..3704)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3750..3805
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3804..3823)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(4164..4180)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4164..4180)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4177..4199)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4188..4204
                     /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(4212..4218,4219..4236,4237..4242))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4257..4278
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4337..4532
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4383..4518
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4538..4672
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4575..4594)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4629..4648
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4740..4757)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4911..5499)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4991..5010)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5670..6461,6462..6530))
                     /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     6293..6312
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6531..6635)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
       61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      181 ggactttcca ttgacgtcaa tgggtggact atttacggta aactgcccac ttggcagtac
      241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      361 tattagtcat cgctattacc atgggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggcgcgc gccaggcggg gcggggcggg gcgaggggcg
      541 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgttgcctt cgccccgtgc cccgctccgc gccgcctcgc gccgcccgcc
      721 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc
      781 gggctgtaat tagcgcttgg tttaatgacg gctcgtttct tttctgtggc tgcgtgaaag
      841 ccttaaaggg ctccgggagg gccctttgtg cgggggggag cggctcgggg ggtgcgtgcg
      901 tgtgtgtgtg cgtggggagc gccgcgtgcg gcccgcgctg cccggcggct gtgagcgctg
      961 cgggcgcggc gcggggcttt gtgcgctccg cgtgtgcgcg aggggagcgc ggccgggggc
     1021 ggtgccccgc ggtgcggggg ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg
     1081 tgggggggtg agcagggggt gtgggcgcgg cggtcgggct gtaacccccc cctgcacccc
     1141 cctccccgag ttgctgagca cggcccggct tcgggtgcgg ggctccgtgc ggggcgtggc
     1201 gcggggctcg ccgtgccggg cggggggtgg cggcaggtgg gggtgccggg cggggcgggg
     1261 ccgcctcggg ccggggaggg ctcgggggag gggcgcggcg gccccggagc gccggcggct
     1321 gtcgaggcgc ggcgagccgc agccattgcc ttttatggta atcgtgcgag agggcgcagg
     1381 gacttccttt gtcccaaatc tggcggagcc gaaatctggg aggcgccgcc gcaccccctc
     1441 tagcgggcgc gggcgaagcg gtgcggcgcc ggcaggaagg aaatgggcgg ggagggcctt
     1501 cgtgcgtcgc cgcgccgccg tccccttctc catctccagc ctcggggctg ccgcaggggg
     1561 acggctgcct tcggggggga cggggcaggg cggggttcgg cttctggcgt gtgaccggcg
     1621 gctctagagc ctctgctaac catgttcatg ccttcttctt tttcctacag ctcctgggca
     1681 acgtgctggt tattgtgctg tctcatcatt ttggcaaaga attcatggtg agcaagggcg
     1741 aggaggataa catggccatc atcaaggagt tcatgcgctt caaggtgcac atggagggct
     1801 ccgtgaacgg ccacgagttc gagatcgagg gcgagggcga gggccgcccc tacgagggca
     1861 cccagaccgc caagctgaag gtgaccaagg gtggccccct gcccttcgcc tgggacatcc
     1921 tgtcccctca gttcatgtac ggctccaagg cctacgtgaa gcaccccgcc gacatccccg
     1981 actacttgaa gctgtccttc cccgagggct tcaagtggga gcgcgtgatg aacttcgagg
     2041 acggcggcgt ggtgaccgtg acccaggact cctccctgca ggacggcgag ttcatctaca
     2101 aggtgaagct gcgcggcacc aacttcccct ccgacggccc cgtaatgcag aagaagacca
     2161 tgggctggga ggcctcctcc gagcggatgt accccgagga cggcgccctg aagggcgaga
     2221 tcaagcagag gctgaagctg aaggacggcg gccactacga cgctgaggtc aagaccacct
     2281 acaaggccaa gaagcccgtg cagctgcccg gcgcctacaa cgtcaacatc aagttggaca
     2341 tcacctccca caacgaggac tacaccatcg tggaacagta cgaacgcgcc gagggccgcc
     2401 actccaccgg cggcatggac gagctgtaca agagtgctgg tggtggagtg caggtggaaa
     2461 ccatctcccc aggagacggg cgcaccttcc ccaagcgcgg ccagacctgc gtggtgcact
     2521 acaccgggat gcttgaagat ggaaagaaat ttgattcctc ccgggacaga aacaagccct
     2581 ttaagtttat gctaggcaag caggaggtga tccgaggctg ggaagaaggg gttgcccaga
     2641 tgagtgtggg tcagagagcc aaactgacta tatctccaga ttatgcctat ggtgccactg
     2701 ggcacccagg catcatccca ccacatgcca ctctcgtctt cgatgtggag cttctaaaac
     2761 tggaaagtgc tggtggtagt gctggtggta gtgctggtgg tagtgctggt ggtagtgctg
     2821 gtggtcctcg aggtgcggcc gccgaggggg agccggggcg gtcgggactg gagcttgaac
     2881 cagaagagcc tcccggctgg cgggaactcg tccccccaga caccctgcac agcctgccca
     2941 agagccaggt gaagcggcag gaggtcatca gcgagctgct ggtgacagag gcggcccacg
     3001 tgcgcatgct gcgggtgctg cacgacctct tcttccagcc catggcagaa tgcctgttct
     3061 tccccttgga ggagctgcag aacatcttcc ccagcctgga cgagctcatc gaggtgcatt
     3121 ccctgttcct cgatcgcctg atgaagcgga ggcaggagag tggctacctc atcggggaga
     3181 tcggagacgt gctgctggcc cggtttgatg gtgctgaggg ctcctggttc cagaaaatct
     3241 cctcccgctt ctgcagccgc cagtcatttg ccttagagca gctcaaagcc aagcaacgca
     3301 aggaccctcg gttctgtgcc ttcgtgcagg aagctgagag ccgcccgcgg tgccgccgcc
     3361 tgcagctgaa ggacatgatc cccacggaga tgcagcggct gaccaagtac cccctgctcc
     3421 tgcagagcat cgggcagaac acagaagcgc ccacagaacg ggagaaagtg gagctggcag
     3481 ccgagtgctg ccgggaaatt ctacaccacg tcaaccaagc cgtgcgtgac atggaggacc
     3541 tgctgaggct caaggactat cagcggcgcc tggacttgtc ccaccttcgg cagagctgag
     3601 cggccgcact cctcaggtgc aggctgccta tcagaaggtg gtggctggtg tggccaatgc
     3661 cctggctcac aaataccact gagatctttt tccctctgcc aaaaattatg gggacatcat
     3721 gaagcccctt gagcatctga cttctggcta ataaaggaaa tttattttca ttgcaatagt
     3781 gtgttggaat tttttgtgtc tctcactcgg aaggacatat gggagggcaa atcatttaaa
     3841 acatcagaat gagtatttgg tttagagttt ggcaacatat gccatatgct ggctgccatg
     3901 aacaaaggtg gctataaaga ggtcatcagt atatgaaaca gccccctgct gtccattcct
     3961 tattccatag aaaagccttg acttgaggtt agattttttt tatattttgt tttgtgttat
     4021 ttttttcttt aacatcccta aaattttcct tacatgtttt actagccaga tttttcctcc
     4081 tctcctgact actcccagtc atagctgtcc ctcttctctt atgaagatcc ctcgacctgc
     4141 agcccaagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     4201 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga
     4261 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     4321 tcgtgccagc ggatccgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
     4381 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     4441 tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag
     4501 gaggcttttt tggaggccta ggcttttgca aaaagctaac ttgtttattg cagcttataa
     4561 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     4621 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgga tccgctgcat
     4681 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     4741 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     4801 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     4861 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     4921 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     4981 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     5041 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     5101 tctcaatgct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     5161 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     5221 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     5281 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     5341 tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     5401 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     5461 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     5521 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     5581 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     5641 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     5701 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     5761 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     5821 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     5881 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     5941 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     6001 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     6061 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     6121 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     6181 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     6241 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     6301 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     6361 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     6421 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     6481 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     6541 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     6601 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     6661 g
//
