LOCUS       sequence-434850-        5410 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the loss-of-function mutations C199S of the genetically 
            encoded green fluorescent hydrogen peroxide sensor oROS-G in 
            mamalian cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5410)
  AUTHORS   Lee J.D., Won. W., Kimball K., Wang Y., Yeboah F., Evitts K.M., 
            Neiswanger C., Schattauer S., Rappleye M., Bremner S.B., Chun C., 
            Smith N., Mack D.L., Young J.E., Lee C.J., Chavkin C., Berndt A.
  TITLE     Structure-guided engineering of a fast genetically encoded sensor 
            for real-time H2O2 monitoring
  JOURNAL   bioRxiv 2024.01.31.578117
REFERENCE   2  (bases 1 to 5410)
  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: "bioRxiv 
            2024.01.31.578117"
COMMENT     Sequence Label: pC3.1_CMV_oROS-G_LF(C199S)
FEATURES             Location/Qualifiers
     source          1..5410
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        23..380
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        23..214
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(46..66)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      231..366
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             415..1209
                     /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             415..1209
                     /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(469..488)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1079..1098
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(1397..1416)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    1441..1488
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      1817..2405
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2306..2325
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        2580..2883
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2884..3087
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3038..3058
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      3117..3126
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     3737..3758
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3864..3885)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4125..4144)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     polyA_signal    4717..4838
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4754..4773)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4808..4827
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4845..5300)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4982..5003)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5194..5213
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        join(5327..5410,1..21)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 taatattgaa aaaggaagag tcctgaggcg gaaagaacca gctgtggaat gtgtgtcagt
       61 tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
      121 attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
      181 gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc
      241 taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg
      301 cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga ggcttttttg
      361 gaggcctagg cttttgcaaa gatcgatcaa gagacaggat gaggatcgtt tcgcatgatt
      421 gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat
      481 gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag
      541 gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaagac
      601 gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac
      661 gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc
      721 ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg
      781 ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag
      841 cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat
      901 caggggctcg cgccagccga actgttcgcc aggctcaagg cgagcatgcc cgacggcgag
      961 gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc
     1021 ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg
     1081 ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg
     1141 ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag
     1201 ttcttctgag cgggactctg gggttcgaaa tgaccgacca agcgacgccc aacctgccat
     1261 cacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc
     1321 gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc
     1381 ctagggggag gctaactgaa acacggaagg agacaatacc ggaaggaacc cgcgctatga
     1441 cggcaataaa aagacagaat aaaacgcacg gtgttgggtc gtttgttcat aaacgcgggg
     1501 ttcggtccca gggctggcac tctgtcgata ccccaccgag accccattgg ggccaatacg
     1561 cccgcgtttc ttccttttcc ccaccccacc ccccaagttc gggtgaaggc ccagggctcg
     1621 cagccaacgt cggggcggca ggccctgcca tagcctcagg ttactcatat atactttaga
     1681 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
     1741 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     1801 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     1861 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     1921 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     1981 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     2041 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     2101 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     2161 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     2221 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     2281 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     2341 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     2401 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     2461 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gccatgcatt
     2521 agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc
     2581 gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg
     2641 acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa
     2701 tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca
     2761 agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac
     2821 atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc
     2881 atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga
     2941 tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg
     3001 gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta
     3061 cggtgggagg tctatataag cagagctggt ttagtgaacc gtcagatccg ctagccgcca
     3121 ccatggagat ggcgagccag cagggcgaga cgatgtccgg accgctgcac attggtttga
     3181 ttcccacagt tggaccgtac ctgctaccgc atattatccc tatgctgcac cagacctttc
     3241 caaagctgga aatgtatctg catgaagcac agacccacca gttactggcg caactggaca
     3301 gcggcaaact cgattgcgtg atcctcgcac tggtgaaaga gagcgaagca ttcattgaag
     3361 tgccgttgtt tgatgagcca atgttgctgg ctatctatga agatcacccg tgggcgaacc
     3421 gcgaatgcgt accgatggcc gatctggcag gggaaaaact gctgatgctg gaagatggtc
     3481 acagtttgcg cgatcaggca atgggtttct gttttgaagc ctcactcgag aacgtctata
     3541 tcaaggccga caagcagaag aacggcatca aggcgaactt caagatccgc cacaacatcg
     3601 aggacggcgg cgtgcagctc gcctaccact accagcagaa cacccccatc ggcgacggcc
     3661 ccgtgctgct gcccgacaac cactacctga gcgtgcagtc caaactttcg aaagacccca
     3721 acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg
     3781 gcatggacga gctgtacaag ggcggtaccg gagggagcat ggtgagcaag ggcgaggagc
     3841 tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt
     3901 tcagcgtgtc cggcgagggt gagggcgatg ccacctacgg caagctgacc ctgaagttca
     3961 tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg
     4021 gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg
     4081 ccatgcccga aggctacatc caggagcgca ccatcttctt caaggacgac ggcaactaca
     4141 agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg
     4201 gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac aacctgccgg
     4261 accaactggg ggcggattac gatacacact tccgcgcgac cagcctggaa actctgcgca
     4321 acatggtggc ggcaggtagc gggatcactt tactgccagc gctggctgtg ccgccggagc
     4381 gcaaacgcga tggggttgtt tatctgccgt gcattaagcc ggaaccacgc cgcactattg
     4441 gtctggttta tcgtcctggc tcaccgctgc gcagccgcta tgagcagctg gcagaggcca
     4501 tccgcgcaag aatggatggc catttcgata aagttttaaa acaggcggtt taatgagaat
     4561 tctgcagtcg acggtaccgc gggcccggga tccaccggat ctagataact gatcataatc
     4621 agccatacca catttgtaga ggttttactt gctttaaaaa acctcccaca cctccccctg
     4681 aacctgaaac ataaaatgaa tgcaattgtt gttgttaact tgtttattgc agcttataat
     4741 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     4801 tctagttgtg gtttgtccaa actcatcaat gtatcttaag gcgtaaattg taagcgttaa
     4861 tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta accaataggc
     4921 cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt tgagtgttgt
     4981 tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca aagggcgaaa
     5041 aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa gttttttggg
     5101 gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat ttagagcttg
     5161 acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc
     5221 tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa
     5281 tgcgccgcta cagggcgcgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     5341 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     5401 aatgcttcaa
//
