LOCUS       sequence-292559-        5399 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  FRET Standard.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5399)
  AUTHORS   Day RN, Day KH, Pavalko FM
  TITLE     Direct visualization by FRET-FLIM of a putative mechanosome complex 
            involving Src, Pyk2 and MBD2 in living MLO-Y4 cells.
  JOURNAL   PLoS One. 2021 Dec 23;16(12):e0261660. doi: 
            10.1371/journal.pone.0261660. eCollection 2021.
  PUBMED    34941939
REFERENCE   2  (bases 1 to 5399)
  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: "PLoS One.";
            date: "2021-12-23"; volume: "16(12):e0261660. doi"; pages: " 
            10.1371/journal.pone.0261660. eCollection 2021"
COMMENT     Sequence Label: Scarlet I-FRET 10-Venus
FEATURES             Location/Qualifiers
     source          1..5399
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        74..377
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        378..581
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     532..552
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      620..629
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             626..1321
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet-I
                     /note="matures faster than mScarlet"
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFIKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(759..777)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1027..1046
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     regulatory      1352..1361
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1358..1360,1361..1363,1364..2077)
                     /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"
     primer_bind     complement(1403..1424)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1664..1683)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2011..2032
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2200..2321
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2237..2256)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2291..2310
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2328..2783)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2465..2486)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2677..2696
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2810..2914
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2916..3273
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2916..3107
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2939..2959)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3124..3259
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3308..4102
                     /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             3308..4102
                     /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(3362..3381)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3972..3991
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4290..4309)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4334..4381
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4710..5298
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5199..5218
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 taccgccatg cattagttat taatagtaat caattacggg gtcattagtt catagcccat
       61 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
      121 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
      181 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
      241 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
      301 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
      361 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
      421 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
      481 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
      541 gcggtaggcg tgtacggtgg gaggtctata taagcagagc tggtttagtg aaccgtcaga
      601 tccgctagcg ctaccggtcg ccaccatggt gagcaagggc gaggcagtga tcaaggagtt
      661 catgcggttc aaggtgcaca tggagggctc catgaacggc cacgagttcg agatcgaggg
      721 cgagggcgag ggccgcccct acgagggcac ccagaccgcc aagctgaagg tgaccaaggg
      781 tggccccctg cccttctcct gggacatcct gtcccctcag ttcatgtacg gctccagggc
      841 cttcatcaag caccccgccg acatccccga ctactataag cagtccttcc ccgagggctt
      901 caagtgggag cgcgtgatga acttcgagga cggcggcgcc gtgaccgtga cccaggacac
      961 ctccctggag gacggcaccc tgatctacaa ggtgaagctc cgcggcacca acttccctcc
     1021 tgacggcccc gtaatgcaga agaagacaat gggctgggaa gcgtccaccg agcggttgta
     1081 ccccgaggac ggcgtgctga agggcgacat taagatggcc ctgcgcctga aggacggcgg
     1141 ccgctacctg gcggacttca agaccaccta caaggccaag aagcccgtgc agatgcccgg
     1201 cgcctacaac gtcgaccgca agttggacat cacctcccac aacgaggact acaccgtggt
     1261 ggaacagtac gaacgctccg agggccgcca ctccaccggc ggcatggacg agctgtacaa
     1321 gtccggactc agatcttcgg atccaccggt cgccaccatg gtgagcaagg gcgaggagct
     1381 gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt
     1441 cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc aagctgaccc tgaagctgat
     1501 ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgggctacgg
     1561 cctgcagtgc ttcgcccgct accccgacca catgaagcag cacgacttct tcaagtccgc
     1621 catgcccgaa ggctacgtcc aggagcgcac catcttcttc aaggacgacg gcaactacaa
     1681 gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg
     1741 catcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca actacaacag
     1801 ccacaacgtc tatatcaccg ccgacaagca gaagaacggc atcaaggcca acttcaagat
     1861 ccgccacaac atcgaggacg gcggcgtgca gctcgccgac cactaccagc agaacacccc
     1921 catcggcgac ggccccgtgc tgctgcccga caaccactac ctgagctacc agtccaagct
     1981 gagcaaagac cccaacgaga agcgcgatca catggtcctg ctggagttcg tgaccgccgc
     2041 cgggatcact ctcggcatgg acgagctgta caagtaaagc ggccgcgact ctagatcata
     2101 atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc
     2161 ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat
     2221 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     2281 cattctagtt gtggtttgtc caaactcatc aatgtatctt aaggcgtaaa ttgtaagcgt
     2341 taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata
     2401 ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt
     2461 tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg
     2521 aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt
     2581 ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc
     2641 ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg
     2701 cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct
     2761 taatgcgccg ctacagggcg cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     2821 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     2881 ataaatgctt caataatatt gaaaaaggaa gagtcctgag gcggaaagaa ccagctgtgg
     2941 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     3001 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     3061 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     3121 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     3181 ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga
     3241 ggaggctttt ttggaggcct aggcttttgc aaagatcgat caagagacag gatgaggatc
     3301 gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag
     3361 gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg
     3421 gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa
     3481 tgaactgcaa gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc
     3541 agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc
     3601 ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga
     3661 tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa
     3721 acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct
     3781 ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgagcat
     3841 gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt
     3901 ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta
     3961 tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga
     4021 ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg
     4081 ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg
     4141 cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
     4201 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     4261 ttcttcgccc accctagggg gaggctaact gaaacacgga aggagacaat accggaagga
     4321 acccgcgcta tgacggcaat aaaaagacag aataaaacgc acggtgttgg gtcgtttgtt
     4381 cataaacgcg gggttcggtc ccagggctgg cactctgtcg ataccccacc gagaccccat
     4441 tggggccaat acgcccgcgt ttcttccttt tccccacccc accccccaag ttcgggtgaa
     4501 ggcccagggc tcgcagccaa cgtcggggcg gcaggccctg ccatagcctc aggttactca
     4561 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     4621 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     4681 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     4741 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     4801 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt
     4861 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     4921 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     4981 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     5041 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     5101 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     5161 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     5221 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     5281 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     5341 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtat
//
