LOCUS       sequence-310121-        5301 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  MPAct with increased linker flexiblity.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5301)
  AUTHORS   Bisaria A, Hayer A, Garbett D, Cohen D, Meyer T
  TITLE     Membrane-proximal F-actin restricts local membrane protrusions and 
            directs cell migration.
  JOURNAL   Science. 2020 Jun 12;368(6496):1205-1210. doi: 
            10.1126/science.aay7794.
  PUBMED    32527825
REFERENCE   2  (bases 1 to 5301)
  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: "10.1126/science"; 
            journalName: "Science"; date: "2020-06-12- 12"; volume: "368"; 
            issue: "6496"; pages: "1205-1210"
COMMENT     Sequence Label: C1-FT-mCitrine-EEVEE-CaaX
FEATURES             Location/Qualifiers
     source          1..5301
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      778..787
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(784..786,787..789,790..1500)
                     /codon_start=1
                     /product="enhanced YFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(829..850)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1090..1109)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1437..1458
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1546..1893
                     /codon_start=1
                     /product="long flexible linker consisting of SAGG repeats 
                     (Komatsu et al., 2011)"
                     /label=EV linker
                     /translation="SAGGSAGGSAGGSAGGSAGGSGSAGGSAGGSTSAGGSAGGSAGGS
                     AGGSAGGSGSAGGSAGGSTSAGGSAGGSAGGSAGGSAGGSGSAGGSAGGSTSAGGSAGG
                     SAGGSAGGSAGG"
     polyA_signal    2089..2210
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2126..2145)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2180..2199
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2217..2672)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2354..2375)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2566..2585
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2699..2803
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2805..3162
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2805..2996
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2828..2848)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3013..3148
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3197..3991
                     /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             3197..3991
                     /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(3251..3270)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3861..3880
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4179..4198)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4223..4270
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4599..5187
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5088..5107
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctcgagct caagcttatg ggcatggcgc gaccacgggg cgcggggccc
      661 tgcagccccg ggttggagcg ggctccgcgc cggagcgtcg gggagctgcg cctgctcttc
      721 gaagcgcgct gcgccgcagt cgccgccgca gcggccgcag ggtcggatcc accggtcgcc
      781 accatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg
      841 gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc
      901 tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc
      961 accctcgtga ccaccttcgg ctacggcctg atgtgcttcg cccgctaccc cgaccacatg
     1021 aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc
     1081 ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc
     1141 ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg
     1201 cacaagctgg agtacaacta caacagccac aacgtctata tcatggccga caagcagaag
     1261 aacggcatca aggtgaactt caagatccgc cacaacatcg aggacggcag cgtgcagctc
     1321 gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac
     1381 cactacctga gctaccagtc caagctgagc aaagacccca acgagaagcg cgatcacatg
     1441 gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag
     1501 agatctcgag ctcaagcttc gaccggtggt ggatccggag gaaccagtgc tggtggtagt
     1561 gctggtggta gtgctggtgg tagtgctggt ggtagtgctg gtggttccgg cagtgctggt
     1621 ggtagtgctg gtggtagtac cagtgctggt ggtagtgctg gtggtagtgc tggtggtagt
     1681 gctggtggta gtgctggtgg ttccggcagt gctggtggta gtgctggtgg tagtaccagt
     1741 gctggtggta gtgctggtgg tagtgctggt ggtagtgctg gtggtagtgc tggtggttcc
     1801 ggcagtgctg gtggtagtgc tggtggtagt accagtgctg gtggtagtgc tggtggtagt
     1861 gctggtggta gtgctggtgg tagtgctggt ggtaccatgg ctgccaattc tggtaaaaag
     1921 aagaaaaaga agtcaaagac aaagtgtgta attatgtaga gatccaccgg atctagataa
     1981 ctgatcataa tcagccatac cacatttgta gaggttttac ttgctttaaa aaacctccca
     2041 cacctccccc tgaacctgaa acataaaatg aatgcaattg ttgttgttaa cttgtttatt
     2101 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     2161 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta acgcgtaaat
     2221 tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt
     2281 taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg
     2341 gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt
     2401 caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc
     2461 aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg
     2521 atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa
     2581 aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc
     2641 cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg
     2701 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     2761 ataaccctga taaatgcttc aataatattg aaaaaggaag agtcctgagg cggaaagaac
     2821 cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
     2881 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
     2941 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
     3001 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     3061 tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag
     3121 aagtagtgag gaggcttttt tggaggccta ggcttttgca aagatcgatc aagagacagg
     3181 atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg
     3241 ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc
     3301 cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg
     3361 tgccctgaat gaactgcaag acgaggcagc gcggctatcg tggctggcca cgacgggcgt
     3421 tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg
     3481 cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat
     3541 catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca
     3601 ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca
     3661 ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa
     3721 ggcgagcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa
     3781 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc
     3841 ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga
     3901 atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc
     3961 cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac
     4021 caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg
     4081 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     4141 atgctggagt tcttcgccca ccctaggggg aggctaactg aaacacggaa ggagacaata
     4201 ccggaaggaa cccgcgctat gacggcaata aaaagacaga ataaaacgca cggtgttggg
     4261 tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc actctgtcga taccccaccg
     4321 agaccccatt ggggccaata cgcccgcgtt tcttcctttt ccccacccca ccccccaagt
     4381 tcgggtgaag gcccagggct cgcagccaac gtcggggcgg caggccctgc catagcctca
     4441 ggttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     4501 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     4561 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     4621 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     4681 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     4741 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     4801 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     4861 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     4921 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     4981 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     5041 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     5101 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     5161 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     5221 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     5281 aaccgtatta ccgccatgca t
//
