LOCUS       sequence-180750-        6425 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses mEos3 fusion with dnRac1 (T17N)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6425)
  AUTHORS   Das S, Yin T, Yang Q, Zhang J, Wu YI, Yu J
  TITLE     Single-molecule tracking of small GTPase Rac1 uncovers spatial 
            regulation of membrane translocation and mechanism for polarized 
            signaling.
  JOURNAL   Proc Natl Acad Sci U S A. 2015 Jan 20;112(3):E267-76. doi: 
            10.1073/pnas.1409667112. Epub 2015 Jan 5.
  PUBMED    25561548
REFERENCE   2  (bases 1 to 6425)
  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: 
            "10.1073/pnas.1409667112"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2015-01-20- 20"; volume: "112"; issue: "3"; pages: "E267-76"
COMMENT     Sequence Label: mEos3-dnRac1
FEATURES             Location/Qualifiers
     source          1..6425
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        306..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     711..733
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     795..814
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     primer_bind     856..875
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        856..874
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    875..899
                     /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        915..1024
                     /label=p10 promoter
                     /note="baculovirus promoter for expression in insect cells"
     CDS             1045..1062
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1072..1101
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     protein_bind    1786..1810
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             join(1831..1878,1879..1881,1882..2409)
                     /codon_start=1
                     /product="dominant negative mutant of human Rac family 
                     small GTPase 1"
                     /label=Rac1 (T17N)
                     /translation="MQAIKCVVVGDGAVGKNCLLISYTTNAFPGEYIPTVFDNYSANVM
                     VDGKPVNLGLWDTAGQEDYDRLRPLSYPQTDVFLICFSLVSPASFENVRAKWYPEVRHH
                     CPNTPIILVGTKLDLRDDKDTIEKLKEKKLTPITYPQGLAMAKEIGAVKYLECSALTQR
                     GLKTVFDEAIRAVLCPPPVKKRKRKCLLL"
     CDS             1831..2409
                     /codon_start=1
                     /product="human Rac family small GTPase 1"
                     /label=Rac1
                     /translation="MQAIKCVVVGDGAVGKNCLLISYTTNAFPGEYIPTVFDNYSANVM
                     VDGKPVNLGLWDTAGQEDYDRLRPLSYPQTDVFLICFSLVSPASFENVRAKWYPEVRHH
                     CPNTPIILVGTKLDLRDDKDTIEKLKEKKLTPITYPQGLAMAKEIGAVKYLECSALTQR
                     GLKTVFDEAIRAVLCPPPVKKRKRKCLLL"
     protein_bind    complement(2426..2450)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             2493..2525
                     /codon_start=1
                     /product="HSV (herpes simplex virus) epitope tag"
                     /label=HSV tag
                     /translation="QPELAPEDPED"
     CDS             2532..2555
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     primer_bind     complement(2645..2664)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2710..2765
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2764..2783)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     terminator      2853..2900
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     misc_recomb     2913..3618
                     /label=baculovirus recombination region (ORF1629)
                     /note="contains part of ORF1629"
     3'UTR           3001..3373
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     CDS             complement(3030..3618)
                     /codon_start=2
                     /product="baculovirus capsid-associated protein"
                     /label=ORF1629
                     /note="required for viral replication"
                     /translation="IPKSSTTNLIADVLADTINRRRVAMAKSSSEATSNDEGWDDDDNR
                     PNKANTPDVKYVQALFNVFTSSQLYTNDSDERNTKAHNILNDVEPLLQNKTQTNIDKAR
                     LLLQDLASFVALSENPLDSPAIGSEKQPLFETNRNLFYKSIEDLIFKFRYKDAENHLIF
                     ALTYHPKDYKFNELLKYVQQLSVNQQRTESSA"
     protein_bind    complement(3627..3660)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     rep_origin      complement(3828..4415)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3907..3926)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4428..5219,5220..5288))
                     /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     5051..5070
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     misc_recomb     5531..6358
                     /label=baculovirus recombination region (lef2/ORF603)
                     /note="contains ORF603 and part of lef2"
     CDS             5531..5655
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(5693..6298)
                     /codon_start=1
                     /product="baculovirus ORF603 protein"
                     /label=ORF603
                     /translation="MAVIFNNKQLLADNSIEKGGELFLFNGSYNILESYVNPVLLKNGV
                     IELEEAAYYAGNILYKTDDPKFIDYINLIIKATHSEELPENSTVVNYRKTMRSGTIHPI
                     KKDIYIYDNKKFTLYDRYIYGYDNNYVNFYEEKNEKEKEYEEEDDKASSLCENKIILSQ
                     INCESFENDFKYYLSDYNYAFSIIDNTTNVLVAFGLYR"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtccgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta cgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catgctgatg cggttttggc agtacaccaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta ataaccccgc cccgttgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagacgtcg tttagtgaac cgtcagatca ctagatgctt
      541 tattgcggta gtttatcaca gttaaattgc taacgccagt ctcgaactta acgtgcagaa
      601 gttggtcgtg aggcactggg caggtaagta tcgggccctt tgtgcggggg gagcggctcg
      661 gggctgtccg cggggggacg gctgccttcg ggggggacgg ggcagggcgg ggttcggctt
      721 ctggcgtgtg accggcggct ctagagcctc tgctaaccat gttcatgcct tcttcttttt
      781 cctacagctc ctgggcaacg tgctggttat tgtgctgtct catcattttg gcaaagaatt
      841 ggatcggacc gaaattaata cgactcacta taggggaatt gtgagcggat aacaattccc
      901 cggagttaat ccgggacctt taattcaacc caacacaata tattatagtt aaataagaat
      961 tattatcaaa tcatttgtat attaattaaa atactatact gtaaattaca ttttatttac
     1021 aatcaaagga gatataccat ggcacaccat caccaccacc acgggagcgg ggagcagaaa
     1081 ctaatatcag aggaggatct tggctcaggt tcaggatctg ggagtagtgc gattaagcca
     1141 gacatgaaga tcaaactccg tatggaaggc aacgtaaacg ggcaccactt tgtgatcgac
     1201 ggagatggta caggcaagcc ttttgaggga aaacagagta tggatcttga agtcaaagag
     1261 ggcggacctc tgccttttgc ctttgatatc ctgaccactg cattccatta cggcaacagg
     1321 gtattcgcca aatatccaga caacatacaa gactatttta agcagtcgtt tcctaagggg
     1381 tattcgtggg aacgaagctt gactttcgaa gacgggggca tttgcaatgc cagaaacgac
     1441 ataacaatgg aaggggacac tttctataat aaagttcgat tttatggtac caactttccc
     1501 gccaatggtc cagttatgca gaagaagacg ctgaaatggg agccctccac tgagaaaatg
     1561 tatgtgcgtg atggagtgct gacgggtgat attgaaatgg ctttgttgct tgaaggaaat
     1621 gcccattacc gatgtgactt cagaactact tacaaagcta aggagaaggg tgtcaagtta
     1681 ccaggcgcac actttgtgga ccactgcatt gagattttaa gccatgacaa agattacaac
     1741 aaggttaagc tgtatgagca tgctgttgct cattctggat catcaacaag tttgtacaaa
     1801 aaagcaggct ccgcggccgc ccccttcacc atgcaggcca tcaagtgtgt ggtggtggga
     1861 gacggagctg taggtaaaaa ttgcctactg atcagttaca caaccaatgc atttcctgga
     1921 gaatatatcc ctactgtctt tgacaattat tctgccaatg ttatggtaga tggaaaaccg
     1981 gtgaatctgg gcttatggga tacagctgga caagaagatt atgacagatt acgcccccta
     2041 tcctatccgc aaacagatgt gttcttaatt tgcttttccc ttgtgagtcc tgcatcattt
     2101 gaaaatgtcc gtgcaaagtg gtatcctgag gtgcggcacc actgtcccaa cactcccatc
     2161 atcctagtgg gaactaaact tgatcttagg gatgataaag acacgatcga gaaactgaag
     2221 gagaagaagc tgactcccat cacctatccg cagggtctag caatggctaa ggagattggt
     2281 gctgtaaaat acctggagtg ctcggcgctc acacagcgag gcctcaagac agtgtttgac
     2341 gaagcgatcc gagcagtcct ctgcccgcct cccgtgaaga agaggaagag aaaatgcctg
     2401 ctgttgtaaa agggtgggcg cgccgaccca gctttcttgt acaaagtggt tgatgatcct
     2461 aagcggccgc acagctgtat acacgtgcaa gccagccaga actcgccccg gaagaccccg
     2521 aggatctcga gcaccaccat caccatcacc atcactaagt gattaacctc aggtgcaggc
     2581 tgcctatcag aaggtggtgg ctggtgtggc caatgccctg gctcacaaat accactgaga
     2641 tcgatctttt tccctctgcc aaaaattatg gggacatcat gaagcccctt gagcatctga
     2701 cttctggcta ataaaggaaa tttattttca ttgcaatagt gtgttggaat tttttgtgtc
     2761 tctcactcgg aaggacatat gggagggcaa atcatttaaa acatcagaat gagtatttgg
     2821 tttagagttt ggcaacatat gcccatatgt aactagcata accccttggg gcctctaaac
     2881 gggtcttgag gggttttttg ctgaaagcat gcggaggaaa ttctccttga agtttccctg
     2941 gtgttcaaag taaaggagtt tgcaccagac gcacctctgt tcactggtcc ggcgtattaa
     3001 aacacgatac attgttatta gtacatttat taagcgctag attctgtgcg ttgttgattt
     3061 acagacaatt gttgtacgta ttttaataat tcattaaatt tataatcttt agggtggtat
     3121 gttagagcga aaatcaaatg attttcagcg tctttatatc tgaatttaaa tattaaatcc
     3181 tcaatagatt tgtaaaatag gtttcgatta gtttcaaaca agggttgttt ttccgaaccg
     3241 atggctggac tatctaatgg attttcgctc aacgccacaa aacttgccaa atcttgtagc
     3301 agcaatctag ctttgtcgat attcgtttgt gttttgtttt gtaataaagg ttcgacgtcg
     3361 ttcaaaatat tatgcgcttt tgtatttctt tcatcactgt cgttagtgta caattgactc
     3421 gacgtaaaca cgttaaatag agcttggaca tatttaacat cgggcgtgtt agctttatta
     3481 ggccgattat cgtcgtcgtc ccaaccctcg tcgttagaag ttgcttccga agacgatttt
     3541 gccatagcca cacgacgcct attaattgtg tcggctaaca cgtccgcgat caaatttgta
     3601 gttgagcttt ttggaattgc gatcgcataa cttcgtatag catacattat acgaagttat
     3661 aagctcggaa cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     3721 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     3781 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     3841 cgccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     3901 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     3961 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     4021 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     4081 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     4141 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     4201 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     4261 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     4321 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     4381 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgtta ccaatgctta
     4441 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     4501 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     4561 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     4621 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     4681 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     4741 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     4801 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     4861 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     4921 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     4981 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     5041 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     5101 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     5161 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     5221 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     5281 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     5341 tccgcgcgtt tcctgcatct tttaatcaaa tcccaagatg tgtataaacg cgccggtatg
     5401 tacaggaaga ggtttatact aaactgttac attgcaaacg tggtttcgtg tgccaagtgt
     5461 gaaaaccgat gtttaatcaa ggctctgacg catttctaca accacgactc caagtgtgtg
     5521 ggtgaagtca tgcatctttt aatcaaatcc caagatgtgt ataaaccacc aaactgccaa
     5581 aaaatgaaaa ctgtcgacaa gctctgtccg tttgctggca actgcaaggg tctcaatcct
     5641 atttgtaatt attgaataat aaaacaatta taaatgtcaa atttgttttt tattaacgat
     5701 acaaaccaaa cgcaacaaga acatttgtag tattatctat aattgaaaac gcgtagttat
     5761 aatcgctgag gtaatattta aaatcatttt caaatgattc acagttaatt tgcgacaata
     5821 taattttatt ttcacataaa ctagacgcct tgtcgtcttc ttcttcgtat tccttctctt
     5881 tttcattttt ctcttcataa aaattaacat agttattatc gtatccatat atgtatctat
     5941 cgtatagagt aaattttttg ttgtcataaa tatatatgtc ttttttaatg gggtgtatag
     6001 taccgctgcg catagttttt ctgtaattta caacagtgct attttctggt agttcttcgg
     6061 agtgtgttgc tttaattatt aaatttatat aatcaatgaa tttgggatcg tcggttttgt
     6121 acaatatgtt gccggcatag tacgcagctt cttctagttc aattacacca ttttttagca
     6181 gcaccggatt aacataactt tccaaaatgt tgtacgaacc gttaaacaaa aacagttcac
     6241 ctcccttttc tatactattg tctgcgagca gttgtttgtt gttaaaaata acagccattg
     6301 taatgagacg cacaaactaa tatcacaaac tggaaatgtc tatcaatata tagttgctga
     6361 tggccggcct attaatagta atcaattacg gggtcattag ttcatagccc atatatggag
     6421 ttccg
//
