LOCUS       sequence-99100-e        8432 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of alpha-actinin with internal sstFRET inserted to 
            measure cytoskeletal stress in vivo.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8432)
  AUTHORS   Meng F, Sachs F
  TITLE     Visualizing dynamic cytoplasmic forces with a compliance-matched 
            FRET sensor.
  JOURNAL   J Cell Sci. 2011 Jan 15;124(Pt 2):261-9. doi: 10.1242/jcs.071928. 
            Epub 2010 Dec 20.
  PUBMED    21172803
REFERENCE   2  (bases 1 to 8432)
  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: "J Cell 
            Sci."; date: "2011-01-15"; volume: "124(Pt 2):261-9. doi"; pages: " 
            10.1242/jcs.071928. Epub 2010 Dec 20"
COMMENT     Sequence Label: PEG-Actinin-M-sstFRET
FEATURES             Location/Qualifiers
     source          1..8432
                     /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      1503..1512
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1509..1511,1512..1514,1515..2225)
                     /codon_start=1
                     /product="Venus YFP with monomerizing A206K mutation (Nagai
                     et al., 2002; Kremers et al., 2006)"
                     /label=mVenus
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2553..2555,2556..2558,2559..3269)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     polyA_signal    5220..5341
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5257..5276)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5311..5330
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5348..5803)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5485..5506)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5697..5716
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5830..5934
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5936..6293
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5936..6127
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5959..5979)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      6144..6279
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             6328..7122
                     /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             6328..7122
                     /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(6382..6401)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6992..7011
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(7310..7329)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    7354..7401
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      7730..8318
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8219..8238
                     /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 gctagcatgg
      601 accattatga ttctcagcaa accaacgatt acatgcagcc agaagaggac tgggaccggg
      661 acctgctcct ggacccggcc tgggagaagc agcagagaaa gacattcacg gcatggtgta
      721 actcccacct ccggaaggcg gggacacaga tcgagaacat cgaagaggac ttccgggatg
      781 gcctgaagct catgctgctg ctggaggtca tctcaggtga acgcttggcc aagccagagc
      841 gaggcaagat gagagtgcac aagatctcca acgtcaacaa ggccctggat ttcatagcca
      901 gcaaaggcgt caaactggtg tccatcggag ccgaagaaat cgtggatggg aatgtgaaga
      961 tgaccctggg catgatctgg accatcatcc tgcgctttgc catccaggac atctccgtgg
     1021 aagagacttc agccaaggaa gggctgctcc tgtggtgtca gagaaagaca gccccttaca
     1081 aaaatgtcaa catccagaac ttccacataa gctggaagga tggcctcggc ttctgtgctt
     1141 tgatccaccg acaccggccc gagctgattg actacgggaa gctgcggaag gatgatccac
     1201 tcacaaatct gaatacggct tttgacgtgg cagagaagta cctggacatc cccaagatgc
     1261 tggatgccga agacatcgtt ggaactgccc gaccggatga gaaagccatc atgacttacg
     1321 tgtctagctt ctaccacgcc ttctctggag cccagaaggc ggagacagca gccaatcgca
     1381 tctgcaaggt gttggccgtc aaccaggaga acgagcagct tatggaagac tacgagaagc
     1441 tggccagtga tctgttggag tggatccgcc gcacaatccc gtggctggag aaccggaccg
     1501 gtgccaccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg
     1561 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg
     1621 ccacctacgg caagctgacc ctgaagctga tctgcaccac cggcaagctg cccgtgccct
     1681 ggcccaccct cgtgaccacc ctgggctacg gcctgcagtg cttcgcccgc taccccgacc
     1741 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca
     1801 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg
     1861 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
     1921 tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcacc gccgacaagc
     1981 agaagaacgg catcaaggcc aacttcaaga tccgccacaa catcgaggac ggcggcgtgc
     2041 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg
     2101 acaaccacta cctgagctac cagtccaaac tgagcaaaga ccccaacgag aagcgcgatc
     2161 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt
     2221 acaagtccgg actcagatct ggaggcttcc acagagatgc tgatgaaacc aaagaatgga
     2281 ttgaagagaa gaatcaagct ctaaacacag acaattatgg acatgatctc gccagtgtcc
     2341 aggccctgca acgcaagcat gagggcttcg agagggacct tgcggctctc ggtgacaagg
     2401 taaactccct tggtgaaaca gcagagcgcc tgatccagtc ccatcccgag tcagcagaag
     2461 acctgcagga aaagtgcaca gagttaaacc aggcctggag cagcctgggg aaacgtgcag
     2521 atcagcgcaa ggcaaaggga ggcgtgaatt ccatggtgag caagggcgag gagctgttca
     2581 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg
     2641 tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca
     2701 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc tggggcgtgc
     2761 agtgcttcgc ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc
     2821 ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc
     2881 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg
     2941 acttcaagga ggacggcaac atcctggggc acaagctgga gtacaacgcc atcagcgaca
     3001 acgtctatat caccgccgac aagcagaaga acggcatcaa ggccaacttc aagatccgcc
     3061 acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg
     3121 gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc aagctgagca
     3181 aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga
     3241 tcactctcgg catggacgag ctgtacaaga agagggcccg ggatccaagc ggccgcgtgc
     3301 ccgagaacac catgcatgcc atgcaacaga agctggagga cttccgggac taccggcgcc
     3361 tgcacaagcc gcccaaggtg caggagaagt gccagctgga gatcaacttc aacacgctgc
     3421 agaccaagct gcggctcagc aaccggcctg ccttcatgcc ctctgagggc aggatggtct
     3481 cggacatcaa caatgcctgg ggctgcctgg agcaggtgga gaagggctat gaggagtggt
     3541 tgctgaatga gatccggagg ctggagcgac tggaccacct ggcagagaag ttccggcaga
     3601 aggcctccat ccacgaggcc tggactgacg gcaaagaggc catgctgcga cagaaggact
     3661 atgagaccgc caccctctcg gagatcaagg ccctgctcaa gaagcatgag gccttcgaga
     3721 gtgacctggc tgcccaccag gaccgtgtgg agcagattgc cgccatcgca caggagctca
     3781 atgagctgga ctattatgac tcacccagtg tcaacgcccg ttgccaaaag atctgtgacc
     3841 agtgggacaa tctgggggcc ctaactcaga agcgaaggga agctctggag cggaccgaga
     3901 aactgctgga gaccattgac cagctgtact tggagtatgc caagcgggct gcacccttca
     3961 acaactggat ggagggggcc atggaggacc tgcaggacac cttcattgtg cacaccattg
     4021 aggagatcca gggactgacc acagcccatg agcagttcaa ggccaccctc cctgatgccg
     4081 acaaggagcg cctggccatc ctgggcatcc acaatgaggt gtccaagatt gtccagacct
     4141 accacgtcaa tatggcgggc accaacccct acacaaccat cacgcctcag gagatcaatg
     4201 gcaaatggga ccacgtgcgg cagctggtgc ctcggaggga ccaagctctg acggaggagc
     4261 atgcccgaca gcagcacaat gagaggctac gcaagcagtt tggagcccag gccaatgtca
     4321 tcgggccctg gatccagacc aagatggagg agatcgggag gatctccatt gagatgcatg
     4381 ggaccctgga ggaccagctc agccacctgc ggcagtatga gaagagcatc gtcaactaca
     4441 agccaaagat tgatcagctg gagggcgacc accagctcat ccaggaggcg ctcatcttcg
     4501 acaacaagca caccaactac accatggagc acatccgtgt gggctgggag cagctgctca
     4561 ccaccatcgc caggaccatc aatgaggtag agaaccagat cctgacccgg gatgccaagg
     4621 gcatcagcca ggagcagatg aatgagttcc gggcctcctt caaccacttt gaccgggatc
     4681 actccggcac actgggtccc gaggagttca aagcctgcct catcagcttg ggttatgata
     4741 ttggcaacga cccccaggga gaagcagaat ttgcccgcat catgagcatt gtggacccca
     4801 accgcctggg ggtagtgaca ttccaggcct tcattgactt catgtcccgc gagacagccg
     4861 acacagatac agcagaccaa gtcatggctt ccttcaagat cctggctggg gacaagaact
     4921 acattaccat ggacgagctg cgccgcgagc tgccacccga ccaggctgag tactgcatcg
     4981 cgcggatggc cccctacacc ggccccgact ccgtgccagg tgctctggac tacatgtcct
     5041 tctccacggc gctgtacggc gagagtgacc tctaaggtac cgcgggcccg ggatccaccg
     5101 gatctagata actgatcata atcagccata ccacatttgt agaggtttta cttgctttaa
     5161 aaaacctccc acacctcccc ctgaacctga aacataaaat gaatgcaatt gttgttgtta
     5221 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
     5281 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     5341 aacgcgtaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc
     5401 agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag
     5461 accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg
     5521 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca
     5581 tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa
     5641 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg
     5701 aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta
     5761 accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtcaggtgg cacttttcgg
     5821 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     5881 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtcctgag
     5941 gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc
     6001 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt
     6061 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     6121 tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc
     6181 cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tcggcctctg
     6241 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaagatcgat
     6301 caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct
     6361 ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc
     6421 tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc
     6481 gacctgtccg gtgccctgaa tgaactgcaa gacgaggcag cgcggctatc gtggctggcc
     6541 acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg
     6601 ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag
     6661 aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc
     6721 ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt
     6781 cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc
     6841 gccaggctca aggcgagcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc
     6901 tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg
     6961 ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag
     7021 cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg
     7081 cagcgcatcg ccttctatcg ccttcttgac gagttcttct gagcgggact ctggggttcg
     7141 aaatgaccga ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct
     7201 tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc
     7261 gcggggatct catgctggag ttcttcgccc accctagggg gaggctaact gaaacacgga
     7321 aggagacaat accggaagga acccgcgcta tgacggcaat aaaaagacag aataaaacgc
     7381 acggtgttgg gtcgtttgtt cataaacgcg gggttcggtc ccagggctgg cactctgtcg
     7441 ataccccacc gagaccccat tggggccaat acgcccgcgt ttcttccttt tccccacccc
     7501 accccccaag ttcgggtgaa ggcccagggc tcgcagccaa cgtcggggcg gcaggccctg
     7561 ccatagcctc aggttactca tatatacttt agattgattt aaaacttcat ttttaattta
     7621 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     7681 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     7741 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     7801 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     7861 agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     7921 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     7981 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     8041 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     8101 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     8161 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     8221 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     8281 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     8341 tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg
     8401 attctgtgga taaccgtatt accgccatgc at
//
