LOCUS       sequence-246108-        5434 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5434)
  AUTHORS   Shcherbakova DM, Hink MA, Joosen L, Gadella TW, Verkhusha VV
  TITLE     An Orange Fluorescent Protein with a Large Stokes Shift for 
            Single-Excitation Multicolor FCCS and FRET Imaging.
  JOURNAL   J Am Chem Soc. 2012 May 9;134(18):7913-23. doi: 10.1021/ja3018972
  PUBMED    22486524
REFERENCE   2  (bases 1 to 5434)
  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"; journalName: 
            "J Am Chem Soc"; date: "2012-05-9- 9"; volume: "134"; issue: "18"; 
            pages: "7913-23"
COMMENT     Sequence Label: pLSSmOrange-mKate2 caspase-3 biosensor
FEATURES             Location/Qualifiers
     source          1..5434
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      616..625
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             622..1329
                     /codon_start=1
                     /product="monomeric orange fluorescent protein with a large
                     Stokes shift (Shcherbakova et al., 2012)"
                     /label=LSSmOrange
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEENNMAIIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEG
                     FQTVKLKVTKGGPLPFAWDILSPQFTYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGMEASSERMYPEDGALK
                     GEDKLRLKLKDGGHYTSEVKTTYKAKKPVQLPGAYIVDIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     1035..1054
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     regulatory      1396..1405
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1402..2100
                     /codon_start=1
                     /product="monomeric far-red fluorescent protein (Shcherbo 
                     et al., 2009)"
                     /label=mKate2
                     /note="mammalian codon-optimized"
                     /translation="MVSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
                     AVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMAL
                     KLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARY
                     CDLPSKLGHR"
     polyA_signal    2222..2343
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2259..2278)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2313..2332
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2350..2805)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2487..2508)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2699..2718
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2832..2936
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2938..3295
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2938..3129
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2961..2981)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3146..3281
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3330..4124
                     /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             3330..4124
                     /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(3384..3403)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3994..4013
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4312..4331)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4356..4403
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4732..5320
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5221..5240
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctcgccac catggtgagc aagggcgagg agaataacat ggccatcatc
      661 aaggagttca tgcgcttcaa ggtgcgcatg gagggctccg tgaacggcca cgagttcgag
      721 atcgagggcg agggcgaggg ccgcccctac gagggctttc agaccgttaa gctgaaggtg
      781 accaagggtg gccccctgcc cttcgcctgg gacatcttgt cccctcagtt cacctacggc
      841 tccaaggcct acgtgaagca ccccgccgac atccccgact acctcaagct gtccttcccc
      901 gagggcttca agtgggagcg cgtgatgaac ttcgaggacg gcggcgtggt gaccgtgact
      961 caggactcct ccctgcagga cggcgagttc atctacaagg tgaagctgcg cggcaccaac
     1021 ttcccctccg acggccccgt aatgcagaag aagaccatgg gcatggaggc ctcctccgag
     1081 cggatgtacc ccgaggacgg cgccctgaag ggcgaggaca agctcaggct gaagctgaag
     1141 gacggcggcc actacacctc cgaggtcaag accacctaca aggccaagaa gcccgtgcag
     1201 ttgcccggcg cctacatcgt cgacatcaag ttggacatca cctcccacaa cgaggactac
     1261 accatcgtgg aacagtacga acgcgccgag ggccgccact ccaccggcgg catggacgag
     1321 ctgtacaagg aattcggtgg ttctggttct gatgaagttg ataagcttgg tggttctggt
     1381 tctggttcac cggtcgccac catggtgagc gagctgatta aggagaacat gcacatgaag
     1441 ctgtacatgg agggcaccgt gaacaaccac cacttcaagt gcacatccga gggcgaaggc
     1501 aagccctacg agggcaccca gaccatgaga atcaaggcgg tcgagggcgg ccctctcccc
     1561 ttcgccttcg acatcctggc taccagcttc atgtacggca gcaaaacctt catcaaccac
     1621 acccagggca tccccgactt ctttaagcag tccttccccg agggcttcac atgggagaga
     1681 gtcaccacat acgaagacgg gggcgtgctg accgctaccc aggacaccag cctccaggac
     1741 ggctgcctca tctacaacgt caagatcaga ggggtgaact tcccatccaa cggccctgtg
     1801 atgcagaaga aaacactcgg ctgggaggcc tccaccgaga ccctgtaccc cgctgacggc
     1861 ggcctggaag gcagagccga catggccctg aagctcgtgg gcgggggcca cctgatctgc
     1921 aacttgaaga ccacatacag atccaagaaa cccgctaaga acctcaagat gcccggcgtc
     1981 tactatgtgg acagaagact ggaaagaatc aaggaggccg acaaagagac ctacgtcgag
     2041 cagcacgagg tggctgtggc cagatactgc gacctcccta gcaaactggg gcacagatga
     2101 gcggccgcga ctctagatca taatcagcca taccacattt gtagaggttt tacttgcttt
     2161 aaaaaacctc ccacacctcc ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt
     2221 taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     2281 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     2341 ttaaggcgta aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa
     2401 tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat
     2461 agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg
     2521 tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac
     2581 catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta
     2641 aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag
     2701 ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg
     2761 taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc
     2821 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     2881 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtcctg
     2941 aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc
     3001 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa
     3061 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     3121 catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc
     3181 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc
     3241 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaagatcg
     3301 atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt
     3361 ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct
     3421 gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga
     3481 ccgacctgtc cggtgccctg aatgaactgc aagacgaggc agcgcggcta tcgtggctgg
     3541 ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact
     3601 ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg
     3661 agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct
     3721 gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg
     3781 gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt
     3841 tcgccaggct caaggcgagc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg
     3901 cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc
     3961 ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag
     4021 agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt
     4081 cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt
     4141 cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc
     4201 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca
     4261 gcgcggggat ctcatgctgg agttcttcgc ccaccctagg gggaggctaa ctgaaacacg
     4321 gaaggagaca ataccggaag gaacccgcgc tatgacggca ataaaaagac agaataaaac
     4381 gcacggtgtt gggtcgtttg ttcataaacg cggggttcgg tcccagggct ggcactctgt
     4441 cgatacccca ccgagacccc attggggcca atacgcccgc gtttcttcct tttccccacc
     4501 ccacccccca agttcgggtg aaggcccagg gctcgcagcc aacgtcgggg cggcaggccc
     4561 tgccatagcc tcaggttact catatatact ttagattgat ttaaaacttc atttttaatt
     4621 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     4681 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     4741 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     4801 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     4861 gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     4921 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     4981 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     5041 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     5101 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     5161 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     5221 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     5281 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     5341 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     5401 tgattctgtg gataaccgta ttaccgccat gcat
//
