LOCUS       sequence-346322-        5171 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Visualization of active Rho.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5171)
  AUTHORS   Mahlandt EK, Palacios Martinez S, Arts JJG, Tol S, van Buul JD, 
            Goedhart J
  TITLE     Opto-RhoGEFs, an optimized optogenetic toolbox to reversibly control
            Rho GTPase activity on a global to subcellular scale, enabling 
            precise control over vascular endothelial barrier strength.
  JOURNAL   Elife. 2023 Jul 14;12:RP84364. doi: 10.7554/eLife.84364.
  PUBMED    37449837
REFERENCE   2  (bases 1 to 5171)
  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"; journalName: "Elife."; 
            date: "2023-07-14"; volume: "12:RP84364. doi"; pages: " 
            10.7554/eLife.84364"
COMMENT     Sequence Label: HaloTag-3xrGBD
FEATURES             Location/Qualifiers
     source          1..5171
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      153..162
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    1959..2080
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1996..2015)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2050..2069
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2087..2542)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2224..2245)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2436..2455
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2569..2673
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2675..3032
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2675..2866
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2698..2718)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2883..3018
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3067..3861
                     /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             3067..3861
                     /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(3121..3140)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3731..3750
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4049..4068)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4093..4140
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4469..5057
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4958..4977
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tgggaggtct atataagcag agctggttta
      121 gtgaaccgtc agatccgcta gcgctaccgg tcgccaccat ggccgagatc ggcaccggct
      181 tccccttcga cccccactac gtggaggtgc tgggcgagcg catgcactac gttgacgtgg
      241 gcccccgcga cggcaccccc gtgctgttcc tgcacggcaa ccccaccagc agctacgtgt
      301 ggcgcaacat catcccccac gtggccccca cccaccgcac tatcgccccc gacctgatcg
      361 gcatgggcaa gagcgacaag cccgacctgg gctacttctt cgacgaccac gtgcgcttca
      421 tggacgcctt catcgaggcc ctgggcctgg aggaggtggt gctggtgatc cacgactggg
      481 gcagcgccct gggcttccac tgggccaagc gcaaccccga gcgcgtgaag ggcatcgcct
      541 tcatggagtt catccgcccc atccccacct gggacgagtg gcccgagttc gcccgcgaga
      601 ccttccaggc cttccgcacc accgacgtgg gccgcaagct gatcatcgac cagaacgtgt
      661 tcatcgaggg caccctgccc atgggcgtgg tgcgccccct gaccgaggtg gagatggacc
      721 actaccgcga gcccttcctg aaccccgtgg accgcgagcc cctgtggcgc ttccccaacg
      781 agctgcccat cgccggcgag cccgccaaca tcgtggccct ggtggaggag tacatggact
      841 ggctgcacca gagccccgtg cccaagctgc tgttctgggg cacccccggc gtgctgatcc
      901 cccccgccga ggccgcccgc ctggccaaga gcctgcccaa cgttaaggcc gtggacatcg
      961 gccccggcct gaacctgctc caggaggaca accccgacct gatcggcagc gagatcgccc
     1021 gctggctgag caccctggag atcagcggcg tgtacatccc cctcgagtcc atcctggagg
     1081 acctcaatat gctctacatc cggcagatgg cactcagcct ggaggacaca gagctgcaga
     1141 ggaaactaga tcatgagatc cggatgaggg atggggcctg caagctgctg gcagcctgct
     1201 cccagcgaga gcaggctctg gaagccacca agagcctgct ggtgtgcaac agccgtattc
     1261 tcagctacat gggtgagctg cagcggcgaa aggaggccca ggtgctggag aagacaggca
     1321 gggtcgagtc catcctggag gacctcaata tgctctacat ccggcagatg gcactcagcc
     1381 tggaggacac agagctgcag aggaaactag atcatgagat ccggatgagg gatggggcct
     1441 gcaagctgct ggcagcctgc tcccagcgag agcaggctct ggaagccacc aagagcctgc
     1501 tggtgtgcaa cagccgtatt ctcagctaca tgggtgagct gcagcggcga aaggaggccc
     1561 aggtgctgga gaagacaggc aggcgtcgag ttatcctgga ggacctcaat atgctctaca
     1621 tccggcagat ggcactcagc ctggaggaca cagagctgca gaggaaacta gatcatgaga
     1681 tccggatgag ggatggggcc tgcaagctgc tggcagcctg ctcccagcga gagcaggctc
     1741 tggaagccac caagagcctg ctggtgtgca acagccgtat tctcagctac atgggtgagc
     1801 tgcagcggcg aaaggaggcc caggtgctgg agaagacagg ctctagataa ctgatcataa
     1861 tcagccatac cacatttgta gaggttttac ttgctttaaa aaacctccca cacctccccc
     1921 tgaacctgaa acataaaatg aatgcaattg ttgttgttaa cttgtttatt gcagcttata
     1981 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     2041 attctagttg tggtttgtcc aaactcatca atgtatctta acgcgtaaat tgtaagcgtt
     2101 aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag
     2161 gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt
     2221 gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga
     2281 aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg
     2341 gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct
     2401 tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc
     2461 gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt
     2521 aatgcgccgc tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg cggaacccct
     2581 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga
     2641 taaatgcttc aataatattg aaaaaggaag agtcctgagg cggaaagaac cagctgtgga
     2701 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     2761 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca
     2821 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
     2881 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     2941 tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag
     3001 gaggcttttt tggaggccta ggcttttgca aagatcgatc aagagacagg atgaggatcg
     3061 tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg
     3121 ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg
     3181 ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat
     3241 gaactgcaag acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca
     3301 gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg
     3361 gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat
     3421 gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa
     3481 catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg
     3541 gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgagcatg
     3601 cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg
     3661 gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat
     3721 caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac
     3781 cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc
     3841 cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc
     3901 ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg
     3961 gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt
     4021 tcttcgccca ccctaggggg aggctaactg aaacacggaa ggagacaata ccggaaggaa
     4081 cccgcgctat gacggcaata aaaagacaga ataaaacgca cggtgttggg tcgtttgttc
     4141 ataaacgcgg ggttcggtcc cagggctggc actctgtcga taccccaccg agaccccatt
     4201 ggggccaata cgcccgcgtt tcttcctttt ccccacccca ccccccaagt tcgggtgaag
     4261 gcccagggct cgcagccaac gtcggggcgg caggccctgc catagcctca ggttactcat
     4321 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc
     4381 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     4441 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     4501 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     4561 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc
     4621 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     4681 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     4741 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     4801 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     4861 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     4921 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     4981 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     5041 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     5101 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
     5161 ccgccatgca t
//
