LOCUS       Exported                6892 bp ds-DNA     circular SYN 30-APR-2021
DEFINITION  Chemogenetic fluorescent H2O2 reporter targeted to the caveolae.
ACCESSION   .
VERSION     .
KEYWORDS    HyPer7.2DAAO-CAV
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6892)
  AUTHORS   Saeedi Saravi SS, Eroglu E, Waldeck-Weiermair M, Sorrentino A, 
            Steinhorn B, Belousov V, Michel T
  TITLE     Differential endothelial signaling responses elicited by 
            chemogenetic H2O2 synthesis.
  JOURNAL   Redox Biol. 2020 Sep;36:101605. doi: 10.1016/j.redox.2020.101605. 
            Epub 2020 Jun 16.
  PUBMED    32590330
REFERENCE   2  (bases 1 to 6892)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Friday, Apr 30, 2021 from SnapGene Viewer 5.1.4.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Redox
            Biol."; date: "2020-09"; pages: "
            10.1016/j.redox.2020.101605. Epub 2020 Jun 16"
FEATURES             Location/Qualifiers
     source          1..6892
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
                     /note="color: #ffffff"
     misc_feature    17..238
                     /label=eNOS 1-74
                     /note="color: #ff00ff"
     misc_feature    257..1693
                     /label=HyPer7.2
                     /note="color: #ffcc00"
     misc_feature    305..307
                     /label=L97V
                     /note="color: #993366"
     misc_feature    380..382
                     /label=M122V
                     /note="color: #999999"
     primer_bind     868..889
                     /label=EGFP-C
                     /note="EGFP, forward primer"
                     /note="color: black; sequence: CATGGTCCTGCTGGAGTTCGTG"
     primer_bind     complement(998..1019)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
                     /note="color: black; sequence: CGTCGCCGTCCAGCTCGACCAG"
     primer_bind     complement(1259..1278)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
                     /note="color: black; sequence: GTCTTGTAGTTGCCGTCGTC"
     misc_feature    1700..2800
                     /label=DAAO
                     /note="color: #a6acb3"
     promoter        complement(2854..2871)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /note="color: #ffffff; direction: LEFT"
     polyA_signal    2876..3010
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
                     /note="color: #a6acb3"
     primer_bind     complement(2900..2919)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
                     /note="color: black; sequence: GTGGTTTGTCCAAACTCATC"
     primer_bind     2954..2973
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
                     /note="color: black; sequence: GAAATTTGTGATGCTATTGC"
     primer_bind     complement(3123..3143)
                     /label=T3
                     /note="T3 promoter, forward primer"
                     /note="color: black; sequence: GCAATTAACCCTCACTAAAGG"
     promoter        complement(3123..3141)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
                     /note="color: #ffffff; direction: LEFT"
     primer_bind     complement(3162..3178)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: LEFT"
     primer_bind     complement(3162..3178)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
                     /note="color: black; sequence: CAGGAAACAGCTATGAC"
     primer_bind     complement(3175..3197)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
                     /note="color: black; sequence: AGCGGATAACAATTTCACACAGG"
     protein_bind    3186..3202
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator 
                     toinhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
                     /note="color: #31849b"
     promoter        complement(3210..3240)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
                     /note="This reverse directional feature has 3 segments:
                        1: 3210 .. 3216 / #ffffff / -10
                        2: 3217 .. 3234 / #ffffff
                        3: 3235 .. 3240 / #ffffff / -35"
     protein_bind    3255..3276
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the 
                     presenceof cAMP."
                     /note="color: #31849b"
     primer_bind     complement(3393..3410)
                     /label=L4440
                     /note="L4440 vector, forward primer"
                     /note="color: black; sequence: AGCGAGTCAGTGAGCGAG"
     rep_origin      complement(3564..4152)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     primer_bind     complement(3644..3663)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
                     /note="color: black; sequence: GGGAAACGCCTGGTATCTTT"
     CDS             complement(4323..5183)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /note="This feature has 2 segments:
                        1: 4323 .. 5114 / #ccffcc
                        2: 5115 .. 5183 / #ccffcc / signal sequence
                     Cleavage site after base 5114"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4946..4965
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
                     /note="color: black; sequence: ATAATACCGCGCCACATAGC"
     promoter        complement(5184..5288)
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: LEFT"
     rep_origin      complement(5314..5769)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; 
                     arrowindicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     primer_bind     complement(5451..5472)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
                     /note="color: black; sequence: GTGGACTCTTGTTCCAAACTGG"
     primer_bind     5663..5682
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
                     /note="color: black; sequence: AGGGAAGAAAGCGAAAGGAG"
     promoter        5847..6824
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian 
                     cytomegalovirusmajor immediate early transcription unit 
                     IE94"
                     /note="color: #ffffff; direction: RIGHT"
     promoter        6859..6877
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
                     /note="color: #ffffff; direction: RIGHT"
     primer_bind     6859..6876
                     /label=SP6
                     /note="SP6 promoter, forward primer"
                     /note="color: black; sequence: ATTTAGGTGACACTATAG"
ORIGIN
        1 tctttttgca ggatccatgg gcaacttgaa gagtgtgggc caggagcccg ggcccccctg
       61 cggcctgggg ctggggctgg gcctcgggct atgcggcaag cagggcccag cctccccggc
      121 acctgagccc agccgggccc ccgcacccgc caccccgcac gcgccagacc acagcccagc
      181 tcccaacagc cccacgctga cccggcctcc ggaggggccc aagttccctc gcgtgaagag
      241 atctgaattc gccaccatgc acctggctaa tgaggagcaa aacgaattgg aaggtgcttt
      301 taaggtgggc ttgatcttta ccgtcgcccc gtacttgctg cctaagttga tcgtcagcct
      361 gcgccgcacc gccccgaagg tgcctttgat gttggaggag aattacaccc acacccttac
      421 cgagtcactg aaacgcggcg acgttgacgc catcattgtg gcagagccgt ttcaagagcc
      481 gggtattgtg actgaacctt tgtatgacga gccgtttttc gttatcgttc cgaagggcca
      541 tagcttcgag gagttggatg ccgttagccc tcgcatgttg ggcgaagaac aagttctgct
      601 tctgaccgag ggcaactgta tgcgtgacca ggtgcttagc tccgccggct tcaacagcaa
      661 caacgtctat atcatggccg acaagcagaa gaacggcatc aaggccaact tcaagatccg
      721 ccacaacgtc gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat
      781 cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcttccagt ccgtcctgag
      841 caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg
      901 gatcactctc ggcatggacg agctgtacaa cgtggatggc ggtagcggtg gcaccggcag
      961 caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt
     1021 aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct
     1081 gaccctgaag ctgatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac
     1141 caccctcagc tacggcctga agtgcttcgc ccgctacccc gaccacatga agcagcacga
     1201 cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga
     1261 cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg
     1321 catcgagctg aagggcatcg gcttcaagga ggacggcaac atcctggggc acaagctgga
     1381 gtacaacggc cccagttgca gcgagcttgc cgccaaacag cgcattcagg gtctcaccaa
     1441 cacccttcaa ggctcaagca tcaataccat ccgccacatg gttgccagtg gcttggcaat
     1501 cagcgtcttg cctgccactg cccttaccga aaacgaccac atgctgttta gcattatccc
     1561 gtttgaaggc accccgccta gccgccgcgt tgtcctggcc tatcgccgca actttgtccg
     1621 ccctaaggcc ctttcagcca tgaaagccgc aattatgcaa agccaactgc atggtgttag
     1681 cttcatctgc gatgctagca gtggtcacag ccagaagagg gtggtcgtgc tgggctccgg
     1741 agtcatcggg ctctctggcg ccctgattct cgctagaaaa ggctactccg tgcatatcct
     1801 ggccagagac ttgcctgagg atgtcagctc ccagaccttc gcaagcccct gggccggagc
     1861 taactggaca ccatttatga ccctgacaga cggacccaga caagccaagt gggaagagtc
     1921 caccttcaag aaatgggtgg agctggttcc cacaggccac gctatgtggc tgaaagggac
     1981 cagaaggttt gcccagaatg aggatggcct cctgggacat tggtacaagg acatcacccc
     2041 taactatagg ccacttccac cttctgaatg tccacctgga gccattggag tgacatacga
     2101 caccctgagc gtccacgctc ccaagtattg cagatacctt gccagagagc tgcagaaact
     2161 tggggctacc ttcgaaagga gaacagtgac tagcctggag caagccttcg atggagctga
     2221 cctcgtggtc aacgccaccg ggctgggagc taagtccatt gccggcatcg atgaccaggc
     2281 tgccgagccc attaggggac agaccgtgct tgtcaaatct ccctgtaaga ggtgcacaat
     2341 ggatagctcc gaccctgctt ctcccgctta tatcattcct agaccaggag gagaagtgat
     2401 ctgtggcgga acctacggag tcggcgattg ggacctgagc gtgaatcccg aaacagtcca
     2461 aaggattctc aagcactgcc tgagacttga ccctaccatt agctccgatg gaactatcga
     2521 gggcattgaa gtgctgagac ataacgttgg cctgagaccc gccaggagag gaggacctag
     2581 ggtggaggct gaaagaatcg tcttgcccct ggaccggacc aagagccctc tctctctggg
     2641 aagagggtcc gcacgggctg caatggagaa ggaagtgacc cttgtccacg cctacggctt
     2701 tagtagcgca gggtatcagc aatcctgggg agccgtggag gacgtggctc agctggtcga
     2761 tgaagccttc caaagatacc acggcgcagc tagggagagc ctgccccccc tggagcggct
     2821 gaccctgtaa aagcttgcct cgagcctcta gaactatagt gagtcgtatt acgtagatcc
     2881 agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa
     2941 atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa
     3001 taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg
     3061 ggaggttttt taattcgcgg ccgcggcgcc aatgcattgg gcccggtacc cagcttttgt
     3121 tccctttagt gagggttaat tgcgcgcttg gcgtaatcat ggtcatagct gtttcctgtg
     3181 tgaaattgtt atccgctcac aattccacac aacatacgag ccgggagcat aaagtgtaaa
     3241 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     3301 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     3361 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     3421 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     3481 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     3541 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     3601 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     3661 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     3721 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     3781 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     3841 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     3901 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     3961 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     4021 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     4081 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     4141 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     4201 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     4261 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     4321 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     4381 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     4441 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     4501 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     4561 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     4621 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     4681 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     4741 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     4801 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     4861 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     4921 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     4981 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     5041 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     5101 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     5161 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     5221 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     5281 gttccgcgca catttccccg aaaagtgcca cctaaattgt aagcgttaat attttgttaa
     5341 aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca
     5401 aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga
     5461 acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc
     5521 agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc
     5581 gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc
     5641 cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg
     5701 caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac
     5761 agggcgcgtc ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg
     5821 cctcttcgct attacgccag tcgaccgcca attcaatatg gcgtatatgg actcatgcca
     5881 attcaatatg gtggatctgg acctgtgcca attcaatatg gcgtatatgg actcgtgcca
     5941 attcaatatg gtggatctgg accccagcca attcaatatg gcggacttgg caccatgcca
     6001 attcaatatg gcggacctgg cactgtgcca actggggagg ggtctacttg gcacggtgcc
     6061 aagtttgagg aggggtcttg gccctgtgcc aagtccgcca tattgaattg gcatggtgcc
     6121 aataatggcg gccatattgg ctatatgcca ggatcaatat ataggcaata tccaatatgg
     6181 ccctatgcca atatggctat tggccaggtt caatactatg tattggccct atgccatata
     6241 gtattccata tatgggtttt cctattgacg tagatagccc ctcccaatgg gcggtcccat
     6301 ataccatata tggggcttcc taataccgcc catagccact cccccattga cgtcaatggt
     6361 ctctatatat ggtctttcct attgacgtca tatgggcggt cctattgacg tatatggcgc
     6421 ctcccccatt gacgtcaatt acggtaaatg gcccgcctgg ctcaatgccc attgacgtca
     6481 ataggaccac ccaccattga cgtcaatggg atggctcatt gcccattcat atccgttctc
     6541 acgcccccta ttgacgtcaa tgacggtaaa tggcccactt ggcagtacat caatatctat
     6601 taatagtaac ttggcaagta cattactatt ggaagtacgc cagggtacat tggcagtact
     6661 cccattgacg tcaatggcgg taaatggccc gcgatggctg ccaagtacat ccccattgac
     6721 gtcaatgggg aggggcaatg acgcaaatgg gcgttccatt gacgtaaatg ggcggtaggc
     6781 gtgcctaatg ggaggtctat ataagcaatg ctcgtttagg gaaccgccat tctgcctggg
     6841 gacgtcggag caagcttgat ttaggtgaca ctatagaata caagctactt gt
//
