LOCUS       sequence-200604-        5395 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Localization: Cytosolic FRET, Excitation: , Emission: .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5395)
  AUTHORS   Davidson Lab
  TITLE     FRET Indicators - Live and Fixed Cells
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5395)
  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: 
            "Unpublished"
COMMENT     Sequence Label: Clover-mRuby2-FRET-10
FEATURES             Location/Qualifiers
     source          1..5395
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      535..544
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             541..1251
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby, with improved photophysical properties (Lam et al., 
                     2012)"
                     /label=mRuby2
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTH
                     MALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAV
                     AKFAGLGGGMDELYK"
     regulatory      1276..1285
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1282..2001
                     /codon_start=1
                     /product="monomeric derivative of the bright green-yellow 
                     fluorescent protein Clover"
                     /label=mClover2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGVACFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIK
                     ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1327..1348)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     1935..1956
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2123..2244
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2160..2179)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2214..2233
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2251..2706)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2388..2409)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2600..2619
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2733..2837
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2839..3196
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2839..3030
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2862..2882)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3047..3182
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3231..4025
                     /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             3231..4025
                     /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(3285..3304)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3895..3914
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4213..4232)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4257..4304
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4633..5221
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5122..5141
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagccacc
      541 atggtgtcta agggcgaaga gctgatcaag gaaaatatgc gtatgaaggt ggtcatggaa
      601 ggttcggtca acggccacca attcaaatgc acaggtgaag gagaaggcaa tccgtacatg
      661 ggaactcaaa ccatgaggat caaagtcatc gagggaggac ccctgccatt tgcctttgac
      721 attcttgcca cgtcgttcat gtatggcagc cgtactttta tcaagtaccc gaaaggcatt
      781 cctgatttct ttaaacagtc ctttcctgag ggttttactt gggaaagagt tacgagatac
      841 gaagatggtg gagtcgtcac cgtcatgcag gacaccagcc ttgaggatgg ctgtctcgtt
      901 taccacgtcc aagtcagagg ggtaaacttt ccctccaatg gtcccgtgat gcagaagaag
      961 accaagggtt gggagcctaa tacagagatg atgtatccag cagatggtgg tctgagggga
     1021 tacactcata tggcactgaa agttgatggt ggtggccatc tgtcttgctc tttcgtaaca
     1081 acttacaggt caaaaaagac cgtcgggaac atcaagatgc ccggtatcca tgccgttgat
     1141 caccgcctgg aaaggttaga ggaaagtgac aatgaaatgt tcgtagtaca acgcgaacac
     1201 gcagttgcca agttcgccgg gcttggtggt gggatggacg agctgtacaa gtccggactc
     1261 agatccccac cggtcgccac catggtgagc aagggcgagg agctgttcac cggggtggtg
     1321 cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt ccgcggcgag
     1381 ggcgagggcg atgccaccaa cggcaagctg accctgaagt tcatctgcac caccggcaag
     1441 ctgcccgtgc cctggcccac cctcgtgacc accttcggct acggcgtggc ctgcttcagc
     1501 cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac
     1561 gtccaggagc gcaccatctc tttcaaggac gacggtacct acaagacccg cgccgaggtg
     1621 aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag
     1681 gacggcaaca tcctggggca caagctggag tacaacttca acagccacaa cgtctatatc
     1741 acggccgaca agcagaagaa cggcatcaag gctaacttca agatccgcca caacgttgag
     1801 gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc
     1861 gtgctgctgc ccgacaacca ctacctgagc catcagtccg ccctgagcaa agaccccaac
     1921 gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc
     1981 atggacgagc tgtacaagta agcggccgcg actctagatc ataatcagcc ataccacatt
     2041 tgtagaggtt ttacttgctt taaaaaacct cccacacctc cccctgaacc tgaaacataa
     2101 aatgaatgca attgttgttg ttaacttgtt tattgcagct tataatggtt acaaataaag
     2161 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt
     2221 gtccaaactc atcaatgtat cttaaggcgt aaattgtaag cgttaatatt ttgttaaaat
     2281 tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa atcggcaaaa
     2341 tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca gtttggaaca
     2401 agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg
     2461 gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg aggtgccgta
     2521 aagcactaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg ggaaagccgg
     2581 cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa
     2641 gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg ccgctacagg
     2701 gcgcgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct
     2761 aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat
     2821 attgaaaaag gaagagtcct gaggcggaaa gaaccagctg tggaatgtgt gtcagttagg
     2881 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     2941 gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     3001 gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac
     3061 tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga
     3121 ggccgaggcc gcctcggcct ctgagctatt ccagaagtag tgaggaggct tttttggagg
     3181 cctaggcttt tgcaaagatc gatcaagaga caggatgagg atcgtttcgc atgattgaac
     3241 aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact
     3301 gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc
     3361 gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg caagacgagg
     3421 cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg
     3481 tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt
     3541 catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc
     3601 atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag
     3661 cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg
     3721 ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgag catgcccgac ggcgaggatc
     3781 tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt
     3841 ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg
     3901 ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt
     3961 acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct
     4021 tctgagcggg actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg
     4081 agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga
     4141 cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccctag
     4201 ggggaggcta actgaaacac ggaaggagac aataccggaa ggaacccgcg ctatgacggc
     4261 aataaaaaga cagaataaaa cgcacggtgt tgggtcgttt gttcataaac gcggggttcg
     4321 gtcccagggc tggcactctg tcgatacccc accgagaccc cattggggcc aatacgcccg
     4381 cgtttcttcc ttttccccac cccacccccc aagttcgggt gaaggcccag ggctcgcagc
     4441 caacgtcggg gcggcaggcc ctgccatagc ctcaggttac tcatatatac tttagattga
     4501 tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat
     4561 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     4621 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     4681 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     4741 ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt
     4801 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     4861 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     4921 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     4981 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     5041 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     5101 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     5161 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     5221 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     5281 gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcca tgcattagtt
     5341 attaatagta atcaattacg gggtcattag ttcatagccc atatatggag ttccg
//
