LOCUS pCS2+HyPer7-NES 5564 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Mammalian expression of cytoplasm targeted ultrasensitive hydrogen
peroxide indicator HyPer7 for optical imaging.
ACCESSION .
VERSION .
KEYWORDS pCS2+HyPer7-NES
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5564)
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 5564)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
//www.snapgene.com
FEATURES Location/Qualifiers
source 1..5564
/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"
primer_bind 1695..1716
/label=EGFP-C
/note="EGFP, forward primer"
primer_bind complement(1825..1846)
/label=EGFP-N
/note="EGFP, reverse primer"
primer_bind complement(2086..2105)
/label=EXFP-R
/note="For distinguishing EGFP variants, reverse primer"
CDS 2527..2553
/codon_start=1
/product="nuclear export signal from the HIV Rev protein
(Fischer et al., 1995)"
/label=NES
/translation="LPPLERLTL"
promoter complement(2572..2589)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
polyA_signal 2594..2728
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(2618..2637)
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind 2672..2691
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind complement(2841..2861)
/label=T3
/note="T3 promoter, forward primer"
promoter complement(2841..2859)
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
primer_bind complement(2880..2896)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(2880..2896)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(2893..2915)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 2904..2920
/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(2928..2958)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 2973..2994
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(3111..3128)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(3282..3870)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(3362..3381)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(4041..4901)
/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 4664..4683
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(4902..5006)
/gene="bla"
/label=AmpR promoter
rep_origin complement(5032..5487)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(5169..5190)
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind 5381..5400
/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 accatgcacc tggctaatga ggagcaaaac gaattggaag gtgcttttaa gctgggcttg
1141 atctttaccg tcgccccgta cttgctgcct aagttgatcg tcagcctgcg ccgcaccgcc
1201 ccgaagatgc ctttgatgtt ggaggagaat tacacccaca cccttaccga gtcactgaaa
1261 cgcggcgacg ttgacgccat cattgtggca gagccgtttc aagagccggg tattgtgact
1321 gaacctttgt atgacgagcc gtttttcgtt atcgttccga agggccatag cttcgaggag
1381 ttggatgccg ttagccctcg catgttgggc gaagaacaag ttctgcttct gaccgagggc
1441 aactgtatgc gtgaccaggt gcttagctcc gccggcttca acagcaacaa cgtctatatc
1501 atggccgaca agcagaagaa cggcatcaag gccaacttca agatccgcca caacgtcgag
1561 gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc
1621 gtgctgctgc ccgacaacca ctacctgagc ttccagtccg tcctgagcaa agaccccaac
1681 gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc
1741 atggacgagc tgtacaacgt ggatggcggt agcggtggca ccggcagcaa gggcgaggag
1801 ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag
1861 ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagctg
1921 atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctcagctac
1981 ggcctgaagt gcttcgcccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc
2041 gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac
2101 aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag
2161 ggcatcggct tcaaggagga cggcaacatc ctggggcaca agctggagta caacggcccc
2221 agttgcagcg agcttgccgc caaacagcgc attcagggtc tcaccaacac ccttcaaggc
2281 tcaagcatca ataccatccg ccacatggtt gccagtggct tggcaatcag cgtcttgcct
2341 gccactgccc ttaccgaaaa cgaccacatg ctgtttagca ttatcccgtt tgaaggcacc
2401 ccgcctagcc gccgcgttgt cctggcctat cgccgcaact ttgtccgccc taaggccctt
2461 tcagccatga aagccgcaat tatgcaaagc caactgcatg gtgttagctt catctgcgat
2521 accggtctgc cccccctgga gcggctgacc ctgtaactcg agcctctaga actatagtga
2581 gtcgtattac gtagatccag acatgataag atacattgat gagtttggac aaaccacaac
2641 tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt
2701 aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca
2761 ggttcagggg gaggtgtggg aggtttttta attcgcggcc gcggcgccaa tgcattgggc
2821 ccggtaccca gcttttgttc cctttagtga gggttaattg cgcgcttggc gtaatcatgg
2881 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
2941 gggagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
3001 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
3061 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
3121 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
3181 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
3241 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
3301 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
3361 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
3421 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
3481 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
3541 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
3601 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
3661 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
3721 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
3781 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
3841 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
3901 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
3961 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
4021 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
4081 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
4141 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
4201 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
4261 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
4321 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
4381 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
4441 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
4501 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
4561 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
4621 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
4681 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
4741 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
4801 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
4861 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
4921 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
4981 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc taaattgtaa
5041 gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc
5101 aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga
5161 gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag
5221 ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt
5281 ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta
5341 gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag
5401 cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg
5461 cgcttaatgc gccgctacag ggcgcgtccc attcgccatt caggctgcgc aactgttggg
5521 aagggcgatc ggtgcgggcc tcttcgctat tacgccagtc gacc
//
