LOCUS       sequence-224796-        7065 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid driving expression of membrane bound KillerRed fluorophore 
            under UAS response element, with a green lens reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7065)
  AUTHORS   Formella I, Svahn AJ, Radford RAW, Don EK, Cole NJ, Hogan A, Lee A, 
            Chung RS, Morsch M
  TITLE     Real-time visualization of oxidative stress-mediated 
            neurodegeneration of individual spinal motor neurons in vivo.
  JOURNAL   Redox Biol. 2018 Aug 23;19:226-234. doi: 
            10.1016/j.redox.2018.08.011.
  PUBMED    30193184
REFERENCE   2  (bases 1 to 7065)
  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: "Redox 
            Biol."; date: "2018-08-23"; volume: "19:226-234. doi"; pages: " 
            10.1016/j.redox.2018.08.011"
COMMENT     Sequence Label: 4xnrUAS:MLS-KillerRed in pTol2pA2-acrys-EGFP-
FEATURES             Location/Qualifiers
     source          1..7065
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..25
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(840..864)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     polyA_signal    866..1000
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     protein_bind    complement(1063..1083)
                     /gene="mutant version of attB"
                     /label=attB3
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     primer_bind     complement(1091..1107)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    1123..1257
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    join(1696..1883,1884..1895)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(1929..1945)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1982..2002)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1982..2000)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2021..2037)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2034..2056)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2045..2061
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(2069..2075,2076..2093,2094..2099))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2114..2135
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2252..2269)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2423..3011)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2503..2522)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3182..3973,3974..4042))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3805..3824
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4043..4147)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4310..4331)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     misc_feature    join(4680..4691,4692..4854)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     polyA_signal    5222..5343
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(join(5466..6179,6180..6182,6183..6185))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(5469..6179,6180..6182,6183..6185))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(5511..5532)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     5860..5879
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     6119..6140
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      6182..6191
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     6833..6849
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6869..6889
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     primer_bind     6896..6912
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 ccaagtttgt acaaaaaagc aggcttacgg gatcgatcca ccggtcgcca ccatgctgtg
       61 ctgtatgaga agaaccaaac aggttgaaaa gaatgatgag gaccaaaaga tctccgaggg
      121 cggccccgcc ctgttccaga gcgacatgac cttcaaaatc ttcatcgacg gcgaggtgaa
      181 cggccagaag ttcaccatcg tggccgacgg cagcagcaag ttcccccacg gcgacttcaa
      241 cgtgcacgcc gtgtgcgaga ccggcaagct gcccatgagc tggaagccca tctgccacct
      301 gatccagtac ggcgagccct tcttcgcccg ctaccccgac ggcatcagcc atttcgccca
      361 ggagtgcttc cccgagggcc tgagcatcga ccgcaccgtg cgcttcgaga acgacggcac
      421 catgaccagc caccacacct acgagctgga cgacacctgc gtggtgagcc gcatcaccgt
      481 gaactgcgac ggcttccagc ccgacggccc catcatgcgc gaccagctgg tggacatcct
      541 gcccaacgag acccacatgt tcccccacgg ccccaacgcc gtgcgccagc tggccttcat
      601 cggcttcacc accgccgacg gcggcctgat gatgggccac ttcgacagca agatgacctt
      661 caacggcagc cgcgccatcg agatccccgg cccacacttc gtgaccatca tcaccaagca
      721 gatgagggac accagcgaca agcgcgacca cgtgtgccag cgcgaggtgg cctacgccca
      781 cagcgtgccc cgcatcacca gcgccatcgg tagcgacgag gattaaagcg gccaattcta
      841 cccagctttc ttgtacaaag tgggggatcc agacatgata agatacattg atgagtttgg
      901 acaaaccaca actagaatgc agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat
      961 tgctttattt gtaaccatta taagctgcaa taaacaagtt aacaacaaca attgcattca
     1021 ttttatgttt caggttcagg gggaggtgtg ggaggttttt tccaacttta ttatacatag
     1081 ttgataattc actggccgtc gttttacggt accatcgatg atgatccaga catgataaga
     1141 tacattgatg agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt
     1201 gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa acaagttaac
     1261 aacaacaatt gcattcattt tatgtttcag gttcaggggg aggtgtggga ggttttttaa
     1321 agcaagtaaa acctctacaa atgtggtatg gctgattatg atcctctaga tcagatctgc
     1381 gaagatacgg ccacgggtgc tcttgatcct gtggctgatt ttggactgtg ctgctcgcag
     1441 ctgctgatga atcacatact tcctccattt tcttccactg attgactgtt ataatttccc
     1501 taatttccag gtcaaggtgc tgtgcattgt ggtaatagat gtgacatgac gtcacttcca
     1561 aaggaccaat gaacatgtct gaccaatttc atataatgtg aaaacgattt tcataggcag
     1621 aataaataac atttaaatta aactgggcat cagcgcaatt caattggttt ggtaatagca
     1681 agggaaaata gaatgaagtg atctccaaaa aataagtact ttttgactgt aaataaaatt
     1741 gtaaggagta aaaagtactt ttttttctaa aaaaatgtaa ttaagtaaaa gtaaaagtat
     1801 tgatttttaa ttgtactcaa gtaaagtaaa aatccccaaa aataatactt aagtacagta
     1861 atcaagtaaa attactcaag tactttacac ctctggttct tgacccccta ccttcagcaa
     1921 gcccagcaga tccactagtt ctagagcggc cgccaccgcg gtggagctcc agcttttgtt
     1981 ccctttagtg agggttaatt gcgcgcttgg cgtaatcatg gtcatagctg tttcctgtgt
     2041 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     2101 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     2161 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     2221 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     2281 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     2341 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     2401 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     2461 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     2521 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     2581 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     2641 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     2701 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     2761 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     2821 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     2881 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     2941 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     3001 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     3061 actcacgtta aggcattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     3121 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     3181 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     3241 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     3301 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     3361 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     3421 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     3481 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     3541 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     3601 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     3661 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     3721 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     3781 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     3841 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     3901 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     3961 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     4021 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     4081 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     4141 ttccgcgcac atttccccga aaagtgccac ctaaattgta agcgttaata ttttgttaaa
     4201 attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa
     4261 aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa
     4321 caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca
     4381 gggcgatggc ccacgctggc taagaactca tcagcctccc cggtccatct acccacgtac
     4441 caatgcacca attggccaca atgacggcta ctacatggtg ccattccttc ctctttatag
     4501 gaatggagac tacctcctgt ccaacaatgc tcttggatac gagtacgcct acctgttgga
     4561 cccaggtcat tgcacaacac cagaaatgcc ctctgatctg caaaagacgt gaatatctgt
     4621 tcagacaccc atatccactc tgttccacac aggtcagagg tttgtccagg agttcttgac
     4681 agaggtgtaa aaagtactca aaaattttac tcaagtgaaa gtacaagtac ttagggaaaa
     4741 ttttactcaa ttaaaagtaa aagtatctgg ctagaatctt acttgagtaa aagtaaaaaa
     4801 gtactccatt aaaattgtac ttgagtatta aggaagtaaa agtaaaagca agaaagaaaa
     4861 ctagagattc ttgtttaagc ttttaatctc aaaaaacatt aaatgaaatg catacaaggt
     4921 tttatcctgc tttagaactg tttgtattta attatcaaac tataagacag acaatctaat
     4981 gccagtacac gctactcaaa gttgtaaaac ctcagattta acttcagtag aagctgattc
     5041 tcaaaattgt tagtgtcaag cctagctctt ttggggctga aaagcaatcc tgcagtgctg
     5101 aaaagcctct cacaggcagc cgatgcggga agaggtgtat tagtcttgat agagaggctg
     5161 caaatagcag gaaacgtgag cagagactcc ctggtgtctg aaacacaggc cagattggcc
     5221 ttaagataca ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt
     5281 atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa
     5341 gttaacaaca acaattgcat tcattttatg tttcaggttc agggggaggt gtgggaggtt
     5401 ttttaaagca agtaaaacct ctacaaatgt ggtatggctg attatgatct agagtcgcgg
     5461 ccgctttact tgtacagctc gtccatgccg agagtgatcc cggcggcggt cacgaactcc
     5521 agcaggacca tgtgatcgcg cttctcgttg gggtctttgc tcagggcgga ctgggtgctc
     5581 aggtagtggt tgtcgggcag cagcacgggg ccgtcgccga tgggggtgtt ctgctggtag
     5641 tggtcggcga gctgcacgct gccgtcctcg atgttgtggc ggatcttgaa gttcaccttg
     5701 atgccgttct tctgcttgtc ggccatgata tagacgttgt ggctgttgta gttgtactcc
     5761 agcttgtgcc ccaggatgtt gccgtcctcc ttgaagtcga tgcccttcag ctcgatgcgg
     5821 ttcaccaggg tgtcgccctc gaacttcacc tcggcgcggg tcttgtagtt gccgtcgtcc
     5881 ttgaagaaga tggtgcgctc ctggacgtag ccttcgggca tggcggactt gaagaagtcg
     5941 tgctgcttca tgtggtcggg gtagcggctg aagcactgca cgccgtaggt cagggtggtc
     6001 acgagggtgg gccagggcac gggcagcttg ccggtggtgc agatgaactt cagggtcagc
     6061 ttgccgtagg tggcatcgcc ctcgccctcg ccggacacgc tgaacttgtg gccgtttacg
     6121 tcgccgtcca gctcgaccag gatgggcacc accccggtga acagctcctc gcccttgctc
     6181 accatggtgg cgaccggtaa tgtcagacct ggtaactcct tactgtaact tattgcacct
     6241 gatgaacaaa agatgtgaac tgaacgcaag taggaggcta atagaggcta gtacagtaca
     6301 aagatcccca aatgatgtgt gccggcctta gtggaggcag acccctggtt atatacctgt
     6361 gctgcagaaa gaggggctta caaatggatc tctctggaat ggcattatct catggtggga
     6421 acagtgttgt cagcagaatc agcggggtgg actctgacct gagaatgaca gtctagcact
     6481 gaatggctca gacaatactc ttgtggccaa ctatcaccgg aaaataagtg cctgccaaaa
     6541 caaaccagac cagatgaagc tacatggctg tagaacgtca gtatttaatg tgttgtggcc
     6601 acttggttca ccagatttaa gagctaattt ttttaaatat cacagataat caatcgcttt
     6661 tttttttctt gactgaaaag cttagactgg aaaattcttt gaaaaagttt gcactacaga
     6721 aaataaatac ataacatcct gcactgaatg cacactatta atagaaaaga gagcattatt
     6781 gtaggaaatg tattgtatat gtacattctg tgcatttcac aattttctcg agcaggaaac
     6841 agctatgacc atgattacgc caagctatca actttgtata gaaaagttgg gccgctctag
     6901 aactagtgga tcccccgggc tgcagcggag tactgtcctc cgagcggagt actgtcctcc
     6961 gagcggagta ctgtcctccg agcggagtac tgtcctccga gtctagaggg tatataatgg
     7021 atcccatcgc gtctcagcct cactttgagc tcctccacac gaatt
//
