LOCUS pCS2+MLS-HyPer7 5720 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Mammalian expression of mitochondrial matrix targeted ultrasensitive
hydrogen peroxide indicator HyPer7 for optical imaging.
ACCESSION .
VERSION .
KEYWORDS pCS2+MLS-HyPer7
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5720)
AUTHORS Pak VV, Ezerina D, Lyublinskaya OG, Pedre B, Tyurin-Kuzmin PA,
Mishina NM, Thauvin M, Young D, Wahni K, Martinez Gache SA,
Demidovich AD, Ermakova YG, Maslova YD, Shokhina AG, Eroglu E, Bilan
DS, Bogeski I, Michel T, Vriz S, Messens J, Belousov VV
TITLE Ultrasensitive Genetically Encoded Indicator for Hydrogen Peroxide
Identifies Roles for the Oxidant in Cell Migration and Mitochondrial
Function.
JOURNAL Cell Metab. 2020 Mar 3;31(3):642-653.e6. doi:
10.1016/j.cmet.2020.02.003.
PUBMED 32130885
REFERENCE 2 (bases 1 to 5720)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..5720
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 1..978
/label=CMV IE94 promoter
/note="enhancer/promoter region of simian cytomegalovirus
major immediate early transcription unit IE94"
promoter 1013..1031
/label=SP6 promoter
/note="promoter for bacteriophage SP6 RNA polymerase"
primer_bind 1013..1030
/label=SP6
/note="SP6 promoter, forward primer"
CDS 1084..1158
/codon_start=1
/product="mitochondrial presequence of human cytochrome c
oxidase subunit VIII (Rizutto et al., 1989)"
/label=COX8 presequence
/translation="MSVLTPLLLRGLTGSARRLPVPRAK"
CDS 1180..1251
/codon_start=1
/product="mitochondrial presequence of human cytochrome c
oxidase subunit VIII (Rizutto et al., 1989)"
/label=COX8 presequence
/translation="SVLTPLLLRGLTGSARRLPVPRAK"
primer_bind 1884..1905
/label=EGFP-C
/note="EGFP, forward primer"
primer_bind complement(2014..2035)
/label=EGFP-N
/note="EGFP, reverse primer"
primer_bind complement(2275..2294)
/label=EXFP-R
/note="For distinguishing EGFP variants, reverse primer"
promoter complement(2728..2745)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
polyA_signal 2750..2884
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(2774..2793)
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind 2828..2847
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind complement(2997..3017)
/label=T3
/note="T3 promoter, forward primer"
promoter complement(2997..3015)
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
primer_bind complement(3036..3052)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(3036..3052)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(3049..3071)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 3060..3076
/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(3084..3114)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 3129..3150
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(3267..3284)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(3438..4026)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(3518..3537)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(4197..5057)
/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 4820..4839
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(5058..5162)
/gene="bla"
/label=AmpR promoter
rep_origin complement(5188..5643)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(5325..5346)
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind 5537..5556
/label=F1ori-R
/note="F1 origin, reverse primer"
ORIGIN
1 gccaattcaa tatggcgtat atggactcat gccaattcaa tatggtggat ctggacctgt
61 gccaattcaa tatggcgtat atggactcgt gccaattcaa tatggtggat ctggacccca
121 gccaattcaa tatggcggac ttggcaccat gccaattcaa tatggcggac ctggcactgt
181 gccaactggg gaggggtcta cttggcacgg tgccaagttt gaggaggggt cttggccctg
241 tgccaagtcc gccatattga attggcatgg tgccaataat ggcggccata ttggctatat
301 gccaggatca atatataggc aatatccaat atggccctat gccaatatgg ctattggcca
361 ggttcaatac tatgtattgg ccctatgcca tatagtattc catatatggg ttttcctatt
421 gacgtagata gcccctccca atgggcggtc ccatatacca tatatggggc ttcctaatac
481 cgcccatagc cactccccca ttgacgtcaa tggtctctat atatggtctt tcctattgac
541 gtcatatggg cggtcctatt gacgtatatg gcgcctcccc cattgacgtc aattacggta
601 aatggcccgc ctggctcaat gcccattgac gtcaatagga ccacccacca ttgacgtcaa
661 tgggatggct cattgcccat tcatatccgt tctcacgccc cctattgacg tcaatgacgg
721 taaatggccc acttggcagt acatcaatat ctattaatag taacttggca agtacattac
781 tattggaagt acgccagggt acattggcag tactcccatt gacgtcaatg gcggtaaatg
841 gcccgcgatg gctgccaagt acatccccat tgacgtcaat ggggaggggc aatgacgcaa
901 atgggcgttc cattgacgta aatgggcggt aggcgtgcct aatgggaggt ctatataagc
961 aatgctcgtt tagggaaccg ccattctgcc tggggacgtc ggagcaagct tgatttaggt
1021 gacactatag aatacaagct acttgttctt tttgcaggat cccatcgatt cgaattcgcc
1081 accatgtccg tcctgacgcc gctgctgctg cggggcttga caggctcggc ccggcggctc
1141 ccagtgccgc gcgccaagat ccattcgttg ggggatctgt ccgtcctgac gccgctgctg
1201 ctgcggggct tgacaggctc ggcccggcgg ctcccagtgc cgcgcgccaa gatccattcg
1261 ttgggggata ccggtcacct ggctaatgag gagcaaaacg aattggaagg tgcttttaag
1321 ctgggcttga tctttaccgt cgccccgtac ttgctgccta agttgatcgt cagcctgcgc
1381 cgcaccgccc cgaagatgcc tttgatgttg gaggagaatt acacccacac ccttaccgag
1441 tcactgaaac gcggcgacgt tgacgccatc attgtggcag agccgtttca agagccgggt
1501 attgtgactg aacctttgta tgacgagccg tttttcgtta tcgttccgaa gggccatagc
1561 ttcgaggagt tggatgccgt tagccctcgc atgttgggcg aagaacaagt tctgcttctg
1621 accgagggca actgtatgcg tgaccaggtg cttagctccg ccggcttcaa cagcaacaac
1681 gtctatatca tggccgacaa gcagaagaac ggcatcaagg ccaacttcaa gatccgccac
1741 aacgtcgagg acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc
1801 gacggccccg tgctgctgcc cgacaaccac tacctgagct tccagtccgt cctgagcaaa
1861 gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc
1921 actctcggca tggacgagct gtacaacgtg gatggcggta gcggtggcac cggcagcaag
1981 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac
2041 ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc
2101 ctgaagctga tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc
2161 ctcagctacg gcctgaagtg cttcgcccgc taccccgacc acatgaagca gcacgacttc
2221 ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac
2281 ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc
2341 gagctgaagg gcatcggctt caaggaggac ggcaacatcc tggggcacaa gctggagtac
2401 aacggcccca gttgcagcga gcttgccgcc aaacagcgca ttcagggtct caccaacacc
2461 cttcaaggct caagcatcaa taccatccgc cacatggttg ccagtggctt ggcaatcagc
2521 gtcttgcctg ccactgccct taccgaaaac gaccacatgc tgtttagcat tatcccgttt
2581 gaaggcaccc cgcctagccg ccgcgttgtc ctggcctatc gccgcaactt tgtccgccct
2641 aaggcccttt cagccatgaa agccgcaatt atgcaaagcc aactgcatgg tgttagcttc
2701 atctgcgatt gactcgagcc tctagaacta tagtgagtcg tattacgtag atccagacat
2761 gataagatac attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt
2821 tatttgtgaa atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca
2881 agttaacaac aacaattgca ttcattttat gtttcaggtt cagggggagg tgtgggaggt
2941 tttttaattc gcggccgcgg cgccaatgca ttgggcccgg tacccagctt ttgttccctt
3001 tagtgagggt taattgcgcg cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat
3061 tgttatccgc tcacaattcc acacaacata cgagccggga gcataaagtg taaagcctgg
3121 ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag
3181 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
3241 ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
3301 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
3361 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
3421 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
3481 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
3541 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
3601 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg
3661 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
3721 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
3781 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
3841 ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct
3901 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
3961 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
4021 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca
4081 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat
4141 taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac
4201 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt
4261 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt
4321 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag
4381 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct
4441 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt
4501 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc
4561 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt
4621 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg
4681 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg
4741 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct
4801 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc
4861 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt
4921 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt
4981 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg
5041 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat
5101 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg
5161 cgcacatttc cccgaaaagt gccacctaaa ttgtaagcgt taatattttg ttaaaattcg
5221 cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc
5281 cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga
5341 gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg
5401 atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag
5461 cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga
5521 acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg
5581 tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg
5641 cgtcccattc gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt
5701 cgctattacg ccagtcgacc
//
