LOCUS       sequence-148222-        6120 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Caspase-3 FRET biosensor, mammalian expression of 
            mAmetrine-DEVD-tdTomato.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6120)
  AUTHORS   Ai HW, Hazelwood KL, Davidson MW, Campbell RE
  TITLE     Fluorescent protein FRET pairs for ratiometric imaging of dual 
            biosensors.
  JOURNAL   Nat Methods. 2008 May;5(5):401-3. doi: 10.1038/nmeth.1207.
  PUBMED    18425137
REFERENCE   2  (bases 1 to 6120)
  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"; doi: "10.1038/nmeth"; 
            journalName: "Nat Methods"; date: "2008-05"; volume: "5"; issue: 
            "5"; pages: "401-3"
COMMENT     Sequence Label: pmAmetrine-DEVD-tdTomato
FEATURES             Location/Qualifiers
     source          1..6120
                     /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      614..623
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(665..686)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     CDS             1355..2785
                     /codon_start=1
                     /product="tandem dimeric (pseudo-monomeric) derivative of 
                     DsRed (Shaner et al., 2004)"
                     /label=tdTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLGHGTGSTGSGSSGTASSEDNNMAVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPY
                     EGTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVM
                     NFEDGGLVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGV
                     LKGEIHQALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYE
                     RSEGRHHLFLYGMDELYK"
     polyA_signal    2915..3036
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2952..2971)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3006..3025
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3043..3498)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3180..3201)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3392..3411
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3525..3629
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3631..3988
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3631..3822
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3654..3674)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3839..3974
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4023..4817
                     /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             4023..4817
                     /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(4077..4096)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4687..4706
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5005..5024)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5049..5096
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5425..6013
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5914..5933
                     /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 agcgctaccg gtcgccacca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc
      661 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgc gcggcgaggg
      721 cgagggcgat gccaccaacg gcaagctgac cctgaagttc atctgcacct ccggcaagct
      781 gcccgtgccc tggcccaccc tcgtgaccac cctgtcttac ggcgtgcagt gcttcgcccg
      841 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt
      901 ccaggagcgc accatctcct tcaaggacga cggcagctac aggacccgcg ccgaggtgaa
      961 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga
     1021 cggcaacatc ctggggcaca agctggagta caacatgaac gtgtgggacg cgtatatcac
     1081 ggccgacaag cagaagaacg gcatcaaagc gaacttcaag atcgagcaca acgtcgagga
     1141 cggcggcgtg cagctcgccg acgcgtacca gcagaacacc cccatcggcg acggctccgt
     1201 gctgctgcct gacaaccact acctgagctt ccagagcaag ctgttcaaag accccaacga
     1261 gcagcgcgat cacatggtcc tgctggagtt cgttaccgcc gccgggatca ctctcggcgg
     1321 taccggcagt ggcagcggag atgaggtcga tggtatggtg agcaagggcg aggaggtcat
     1381 caaagagttc atgcgcttca aggtgcgcat ggagggctcc atgaacggcc acgagttcga
     1441 gatcgagggc gagggcgagg gccgccccta cgagggcacc cagaccgcca agctgaaggt
     1501 gaccaagggc ggccccctgc ccttcgcctg ggacatcctg tccccccagt tcatgtacgg
     1561 ctccaaggcg tacgtgaagc accccgccga catccccgat tacaagaagc tgtccttccc
     1621 cgagggcttc aagtgggagc gcgtgatgaa cttcgaggac ggcggtctgg tgaccgtgac
     1681 ccaggactcc tccctgcagg acggcacgct gatctacaag gtgaagatgc gcggcaccaa
     1741 cttccccccc gacggccccg taatgcagaa gaagaccatg ggctgggagg cctccaccga
     1801 gcgcctgtac ccccgcgacg gcgtgctgaa gggcgagatc caccaggccc tgaagctgaa
     1861 ggacggcggc cactacctgg tggagttcaa gaccatctac atggccaaga agcccgtgca
     1921 actgcccggc tactactacg tggacaccaa gctggacatc acctcccaca acgaggacta
     1981 caccatcgtg gaacagtacg agcgctccga gggccgccac cacctgttcc tggggcatgg
     2041 caccggcagc accggcagcg gcagctccgg caccgcctcc tccgaggaca acaacatggc
     2101 cgtcatcaaa gagttcatgc gcttcaaggt gcgcatggag ggctccatga acggccacga
     2161 gttcgagatc gagggcgagg gcgagggccg cccctacgag ggcacccaga ccgccaagct
     2221 gaaggtgacc aagggcggcc ccctgccctt cgcctgggac atcctgtccc cccagttcat
     2281 gtacggctcc aaggcgtacg tgaagcaccc cgccgacatc cccgattaca agaagctgtc
     2341 cttccccgag ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg gtctggtgac
     2401 cgtgacccag gactcctccc tgcaggacgg cacgctgatc tacaaggtga agatgcgcgg
     2461 caccaacttc ccccccgacg gccccgtaat gcagaagaag accatgggct gggaggcctc
     2521 caccgagcgc ctgtaccccc gcgacggcgt gctgaagggc gagatccacc aggccctgaa
     2581 gctgaaggac ggcggccact acctggtgga gttcaagacc atctacatgg ccaagaagcc
     2641 cgtgcaactg cccggctact actacgtgga caccaagctg gacatcacct cccacaacga
     2701 ggactacacc atcgtggaac agtacgagcg ctccgagggc cgccaccacc tgttcctgta
     2761 cggcatggac gagctgtaca agtaaggatc caccggatct agataactga tcataatcag
     2821 ccataccaca tttgtagagg ttttacttgc tttaaaaaac ctcccacacc tccccctgaa
     2881 cctgaaacat aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg
     2941 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
     3001 tagttgtggt ttgtccaaac tcatcaatgt atcttaaggc gtaaattgta agcgttaata
     3061 ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg
     3121 aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc
     3181 cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa
     3241 ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt
     3301 cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac
     3361 ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta
     3421 gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg
     3481 cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     3541 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     3601 tgcttcaata atattgaaaa aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt
     3661 gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     3721 gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag
     3781 tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat
     3841 cccgccccta actccgccca gttccgccca ttctccgccc catggctgac taattttttt
     3901 tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg
     3961 cttttttgga ggcctaggct tttgcaaaga tcgatcaaga gacaggatga ggatcgtttc
     4021 gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat
     4081 tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt
     4141 cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac
     4201 tgcaagacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg
     4261 tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc
     4321 aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa
     4381 tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc
     4441 gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg
     4501 aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg agcatgcccg
     4561 acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa
     4621 atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg
     4681 acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct
     4741 tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc
     4801 ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa
     4861 cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat
     4921 cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt
     4981 cgcccaccct agggggaggc taactgaaac acggaaggag acaataccgg aaggaacccg
     5041 cgctatgacg gcaataaaaa gacagaataa aacgcacggt gttgggtcgt ttgttcataa
     5101 acgcggggtt cggtcccagg gctggcactc tgtcgatacc ccaccgagac cccattgggg
     5161 ccaatacgcc cgcgtttctt ccttttcccc accccacccc ccaagttcgg gtgaaggccc
     5221 agggctcgca gccaacgtcg gggcggcagg ccctgccata gcctcaggtt actcatatat
     5281 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     5341 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     5401 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     5461 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     5521 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     5581 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     5641 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     5701 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     5761 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     5821 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     5881 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     5941 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     6001 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     6061 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
//
