LOCUS       sequence-292560-        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-Cerulean3
FEATURES             Location/Qualifiers
     source          1..5399
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        76..379
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        380..583
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     534..554
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      622..631
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             628..1323
                     /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(761..779)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1029..1048
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     regulatory      1354..1363
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1360..1362,1363..1365,1366..2079)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAIHGNVYITADKQKNGIK
                     ANFGLNCNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1405..1426)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1666..1685)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2013..2034
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2202..2323
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2239..2258)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2293..2312
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2330..2785)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2467..2488)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2679..2698
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2812..2916
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2918..3275
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2918..3109
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2941..2961)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3126..3261
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3310..4104
                     /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             3310..4104
                     /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(3364..3383)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3974..3993
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4292..4311)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4336..4383
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4712..5300
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5201..5220
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 attaccgcca tgcattagtt attaatagta atcaattacg gggtcattag ttcatagccc
       61 atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa
      121 cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
      181 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
      241 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
      301 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt
      361 agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg
      421 gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg
      481 gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat
      541 gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctggtttag tgaaccgtca
      601 gatccgctag cgctaccggt cgccaccatg gtgagcaagg gcgaggcagt gatcaaggag
      661 ttcatgcggt tcaaggtgca catggagggc tccatgaacg gccacgagtt cgagatcgag
      721 ggcgagggcg agggccgccc ctacgagggc acccagaccg ccaagctgaa ggtgaccaag
      781 ggtggccccc tgcccttctc ctgggacatc ctgtcccctc agttcatgta cggctccagg
      841 gccttcatca agcaccccgc cgacatcccc gactactata agcagtcctt ccccgagggc
      901 ttcaagtggg agcgcgtgat gaacttcgag gacggcggcg ccgtgaccgt gacccaggac
      961 acctccctgg aggacggcac cctgatctac aaggtgaagc tccgcggcac caacttccct
     1021 cctgacggcc ccgtaatgca gaagaagaca atgggctggg aagcgtccac cgagcggttg
     1081 taccccgagg acggcgtgct gaagggcgac attaagatgg ccctgcgcct gaaggacggc
     1141 ggccgctacc tggcggactt caagaccacc tacaaggcca agaagcccgt gcagatgccc
     1201 ggcgcctaca acgtcgaccg caagttggac atcacctccc acaacgagga ctacaccgtg
     1261 gtggaacagt acgaacgctc cgagggccgc cactccaccg gcggcatgga cgagctgtac
     1321 aagtccggac tcagatcttc ggatccaccg gtcgccacca tggtgagcaa gggcgaggag
     1381 ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag
     1441 ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc
     1501 atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgagctgg
     1561 ggcgtgcagt gcttcgcccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc
     1621 gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac
     1681 aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag
     1741 ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta caacgccatc
     1801 cacggcaacg tctatatcac cgccgacaag cagaagaacg gcatcaaggc caacttcggc
     1861 ctcaactgca acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc
     1921 cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccaag
     1981 ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc
     2041 gccgggatca ctctcggcat ggacgagctg tacaagtaaa gcggccgcga ctctagatca
     2101 taatcagcca taccacattt gtagaggttt tacttgcttt aaaaaacctc ccacacctcc
     2161 ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt taacttgttt attgcagctt
     2221 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     2281 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttaaggcgta aattgtaagc
     2341 gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
     2401 taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt
     2461 gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg
     2521 cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt
     2581 ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga
     2641 gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg
     2701 ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg
     2761 cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc ggggaaatgt gcgcggaacc
     2821 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     2881 tgataaatgc ttcaataata ttgaaaaagg aagagtcctg aggcggaaag aaccagctgt
     2941 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     3001 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     3061 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     3121 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     3181 ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt
     3241 gaggaggctt ttttggaggc ctaggctttt gcaaagatcg atcaagagac aggatgagga
     3301 tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag
     3361 aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc
     3421 cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg
     3481 aatgaactgc aagacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc
     3541 gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg
     3601 ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct
     3661 gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg
     3721 aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat
     3781 ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgagc
     3841 atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg
     3901 gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc
     3961 tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct
     4021 gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat
     4081 cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga
     4141 cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct
     4201 tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg
     4261 agttcttcgc ccaccctagg gggaggctaa ctgaaacacg gaaggagaca ataccggaag
     4321 gaacccgcgc tatgacggca ataaaaagac agaataaaac gcacggtgtt gggtcgtttg
     4381 ttcataaacg cggggttcgg tcccagggct ggcactctgt cgatacccca ccgagacccc
     4441 attggggcca atacgcccgc gtttcttcct tttccccacc ccacccccca agttcgggtg
     4501 aaggcccagg gctcgcagcc aacgtcgggg cggcaggccc tgccatagcc tcaggttact
     4561 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     4621 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     4681 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     4741 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     4801 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     4861 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     4921 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     4981 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     5041 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     5101 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     5161 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     5221 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     5281 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     5341 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgt
//
