LOCUS       sequence-310146-        4917 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Monitor changes in the density of membrane-proximal F-actin (MPA).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4917)
  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 4917)
  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-MPAct-mCitrine
FEATURES             Location/Qualifiers
     source          1..4917
                     /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"
     polyA_signal    1705..1826
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1742..1761)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1796..1815
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1833..2288)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1970..1991)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2182..2201
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2315..2419
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2421..2778
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2421..2612
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2444..2464)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2629..2764
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2813..3607
                     /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             2813..3607
                     /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(2867..2886)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3477..3496
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3795..3814)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3839..3886
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4215..4803
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4704..4723
                     /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 gaattctggt aaaaagaaga aaaagaagtc aaagacaaag
     1561 tgtgtaatta tgtagagatc caccggatct agataactga tcataatcag ccataccaca
     1621 tttgtagagg ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat
     1681 aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa
     1741 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     1801 ttgtccaaac tcatcaatgt atcttaacgc gtaaattgta agcgttaata ttttgttaaa
     1861 attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa
     1921 aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa
     1981 caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca
     2041 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg
     2101 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc
     2161 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc
     2221 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca
     2281 gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     2341 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     2401 atattgaaaa aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta
     2461 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     2521 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     2581 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     2641 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     2701 gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     2761 ggcctaggct tttgcaaaga tcgatcaaga gacaggatga ggatcgtttc gcatgattga
     2821 acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga
     2881 ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg
     2941 gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaagacga
     3001 ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt
     3061 tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct
     3121 gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct
     3181 gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg
     3241 agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca
     3301 ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg agcatgcccg acggcgagga
     3361 tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt
     3421 ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt
     3481 ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct
     3541 ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt
     3601 cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca
     3661 cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg
     3721 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccaccct
     3781 agggggaggc taactgaaac acggaaggag acaataccgg aaggaacccg cgctatgacg
     3841 gcaataaaaa gacagaataa aacgcacggt gttgggtcgt ttgttcataa acgcggggtt
     3901 cggtcccagg gctggcactc tgtcgatacc ccaccgagac cccattgggg ccaatacgcc
     3961 cgcgtttctt ccttttcccc accccacccc ccaagttcgg gtgaaggccc agggctcgca
     4021 gccaacgtcg gggcggcagg ccctgccata gcctcaggtt actcatatat actttagatt
     4081 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     4141 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     4201 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     4261 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     4321 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
     4381 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     4441 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     4501 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     4561 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     4621 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     4681 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     4741 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     4801 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     4861 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc catgcat
//
