LOCUS       sequence-127369-        5396 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 5396)
  AUTHORS   Davidson Lab
  TITLE     FRET Indicators - Live and Fixed Cells
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5396)
  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            "Unpublished"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: mPAGFP-mRuby2-FRET-10
FEATURES             Location/Qualifiers
     source          1..5396
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        68..371
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        372..575
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     526..546
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      602..611
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             608..1318
                     /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      1343..1352
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1349..1351,1352..1354,1355..2065)
                     /codon_start=1
                     /product="photoactivatable GFP variant (Patterson et al., 
                     2002)"
                     /label=PA-GFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFSYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1394..1415)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1655..1674)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2002..2023
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2191..2312
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2228..2247)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2282..2301
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2319..2774)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2456..2477)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2668..2687
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2801..2905
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2907..3264
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2907..3098
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2930..2950)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3115..3250
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3299..4093
                     /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             3299..4093
                     /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(3353..3372)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3963..3982
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4281..4300)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4325..4372
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4701..5289
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5190..5209
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 catgcattag ttattaatag taatcaatta cggggtcatt agttcatagc ccatatatgg
       61 agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc
      121 gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg actttccatt
      181 gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc
      241 atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg
      301 cccagtacat gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg
      361 ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact
      421 cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa
      481 atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta
      541 ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt cagatccgct
      601 agccaccatg gtgtctaagg gcgaagagct gatcaaggaa aatatgcgta tgaaggtggt
      661 catggaaggt tcggtcaacg gccaccaatt caaatgcaca ggtgaaggag aaggcaatcc
      721 gtacatggga actcaaacca tgaggatcaa agtcatcgag ggaggacccc tgccatttgc
      781 ctttgacatt cttgccacgt cgttcatgta tggcagccgt acttttatca agtacccgaa
      841 aggcattcct gatttcttta aacagtcctt tcctgagggt tttacttggg aaagagttac
      901 gagatacgaa gatggtggag tcgtcaccgt catgcaggac accagccttg aggatggctg
      961 tctcgtttac cacgtccaag tcagaggggt aaactttccc tccaatggtc ccgtgatgca
     1021 gaagaagacc aagggttggg agcctaatac agagatgatg tatccagcag atggtggtct
     1081 gaggggatac actcatatgg cactgaaagt tgatggtggt ggccatctgt cttgctcttt
     1141 cgtaacaact tacaggtcaa aaaagaccgt cgggaacatc aagatgcccg gtatccatgc
     1201 cgttgatcac cgcctggaaa ggttagagga aagtgacaat gaaatgttcg tagtacaacg
     1261 cgaacacgca gttgccaagt tcgccgggct tggtggtggg atggacgagc tgtacaagtc
     1321 cggactcaga tccccaccgg tcgccaccat ggtgagcaag ggcgaggagc tgttcaccgg
     1381 ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc
     1441 cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac
     1501 cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ttcagctacg gcgtgcagtg
     1561 cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga
     1621 aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc
     1681 cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt
     1741 caaggaggac ggcaacatcc tggggcacaa gctggagtac aactacaaca gccacaacgt
     1801 ctatatcatg gccgacaagc agaagaacgg catcaaggcc aacttcaaga tccgccacaa
     1861 catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga
     1921 cggccccgtg ctgctgcccg acaaccacta cctgagccac cagtccaagc tgagcaaaga
     1981 ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg ccgggatcac
     2041 tctcggcatg gacgagctgt acaagtaaag cggccgcgac tctagatcat aatcagccat
     2101 accacatttg tagaggtttt acttgcttta aaaaacctcc cacacctccc cctgaacctg
     2161 aaacataaaa tgaatgcaat tgttgttgtt aacttgttta ttgcagctta taatggttac
     2221 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt
     2281 tgtggtttgt ccaaactcat caatgtatct taaggcgtaa attgtaagcg ttaatatttt
     2341 gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat aggccgaaat
     2401 cggcaaaatc ccttataaat caaaagaata gaccgagata gggttgagtg ttgttccagt
     2461 ttggaacaag agtccactat taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt
     2521 ctatcagggc gatggcccac tacgtgaacc atcaccctaa tcaagttttt tggggtcgag
     2581 gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc cgatttagag cttgacgggg
     2641 aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc
     2701 gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc
     2761 gctacagggc gcgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     2821 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     2881 tcaataatat tgaaaaagga agagtcctga ggcggaaaga accagctgtg gaatgtgtgt
     2941 cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     3001 ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     3061 caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg
     3121 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt
     3181 tatgcagagg ccgaggccgc ctcggcctct gagctattcc agaagtagtg aggaggcttt
     3241 tttggaggcc taggcttttg caaagatcga tcaagagaca ggatgaggat cgtttcgcat
     3301 gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg
     3361 ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc
     3421 gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca
     3481 agacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct
     3541 cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga
     3601 tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg
     3661 gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat
     3721 cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga
     3781 gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgagca tgcccgacgg
     3841 cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg
     3901 ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat
     3961 agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct
     4021 cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga
     4081 cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg
     4141 ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt
     4201 ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc
     4261 caccctaggg ggaggctaac tgaaacacgg aaggagacaa taccggaagg aacccgcgct
     4321 atgacggcaa taaaaagaca gaataaaacg cacggtgttg ggtcgtttgt tcataaacgc
     4381 ggggttcggt cccagggctg gcactctgtc gataccccac cgagacccca ttggggccaa
     4441 tacgcccgcg tttcttcctt ttccccaccc caccccccaa gttcgggtga aggcccaggg
     4501 ctcgcagcca acgtcggggc ggcaggccct gccatagcct caggttactc atatatactt
     4561 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     4621 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     4681 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     4741 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     4801 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
     4861 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     4921 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     4981 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     5041 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     5101 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     5161 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     5221 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     5281 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     5341 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgc
//
