LOCUS       Exported                6067 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  AAV expression vector with CMV promoter driving expression of the 
            hydrogen peroxide-sensitive fluorescent biosensor HyPer3..
ACCESSION   .
VERSION     .
KEYWORDS    pAAV-HyPer3
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6067)
  AUTHORS   Steinhorn B, Sorrentino A, Badole S, Bogdanova Y, Belousov V, Michel
            T
  TITLE     Chemogenetic generation of hydrogen peroxide in the heart induces 
            severe cardiac dysfunction.
  JOURNAL   Nat Commun. 2018 Oct 2;9(1):4044. doi: 10.1038/s41467-018-06533-2.
  PUBMED    30279532
REFERENCE   2  (bases 1 to 6067)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6067
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     enhancer        206..509
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        510..713
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     664..684
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     710..734
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      1335..1344
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             1341..2777
                     /codon_start=1
                     /product="improved fluorescent indicator for intracellular 
                     H2O2 (Bilan et al., 2013)"
                     /label=HyPer-3
                     /translation="MEMASQQGETMSGPLHIGLIPTVGPYLLPHIIPMLYQTFPKLEMY
                     LHEAQTHQLLAQLDSGKLDCVILALVKESEAFIEVPLFDEPMLLAIYEDHPWANRECVP
                     MADLAGEKLLMLEDGHCLRDQAMSAGYNSDNVYIMADKQKNGIKANFKIRHNVEDGSVQ
                     LADHYQQNTPIGDGPVLLPDNHYLSFQSVLSKDPNEKRDHMVLLEFVTAAGITLGMDEL
                     YNVDGGSGGTGSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKLICT
                     TGKLPVPWPTLVTTLGYGLKCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGNYKT
                     RAEVKFEGDTLVNRIELKGIGFKEDGNILGHKLEYNGTGFCFEAGADEDTHFRATSLET
                     LRNMVAAGSGITLLPALAVPPERKRDGVVYLPCIKPEPRRTIGLVYRPGSPLRSRYEQL
                     AEAIRARMDGHFDKVLKQAV"
     primer_bind     1949..1970
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2079..2100)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2340..2359)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     polyA_signal    2814..3290
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3013..3032)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3330..3470
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3330..3459
                     /label=AAV2 ITR
     rep_origin      3545..4000
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3632..3651)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3842..3863
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4017..4036)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4136..4158
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4196..4214)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4282..4386
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4387..5247
                     /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     complement(4605..4624)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5418..6006
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5907..5926
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtctagtt attaatagta atcaattacg gggtcattag
      181 ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct
      241 gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc
      301 caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg
      361 cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat
      421 ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca
      481 tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc
      541 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga
      601 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat
      661 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctcgtttag
      721 tgaaccgtca gatcgcctgg agacgccatc cacgctgttt tgacctccat agaagacacc
      781 gggaccgatc cagcctccgc ggattcgaat cccggccggg aacggtgcat tggaacgcgg
      841 attccccgtg ccaagagtga cgtaagtacc gcctatagag tctataggcc cacaaaaaat
      901 gctttcttct tttaatatac ttttttgttt atcttatttc taatactttc cctaatctct
      961 ttctttcagg gcaataatga tacaatgtat catgcctctt tgcaccattc taaagaataa
     1021 cagtgataat ttctgggtta aggcaatagc aatatttctg catataaata tttctgcata
     1081 taaattgtaa ctgatgtaag aggtttcata ttgctaatag cagctacaat ccagctacca
     1141 ttctgctttt attttatggt tgggataagg ctggattatt ctgagtccaa gctaggccct
     1201 tttgctaatc atgttcatac ctcttatctt cctcccacag ctcctgggca acgtgctggt
     1261 ctgtgtgctg gcccatcact ttggcaaaga attgggattc gaacatcgat tgaattcccc
     1321 ggggatcctc tagcgccacc atggagatgg caagccagca gggcgagacg atgtccggac
     1381 cgctgcacat tggtttgatt cccacagttg gaccgtacct gctaccgcat attatcccta
     1441 tgctgtacca gacctttcca aagctggaaa tgtatctgca tgaagcacag acccaccagt
     1501 tactggcgca actggacagc ggcaaactcg attgcgtgat cctcgcactg gtgaaagaga
     1561 gcgaagcatt cattgaagtg ccgttgtttg atgagccaat gttgctggct atctatgaag
     1621 atcacccgtg ggcgaaccgc gaatgcgtac cgatggccga tctggcaggg gaaaaactgc
     1681 tgatgctgga agatggtcac tgtttgcgcg atcaggcaat gtccgccggc tacaacagcg
     1741 acaacgtcta tatcatggcc gacaagcaga agaacggcat caaggccaac ttcaagatcc
     1801 gccacaacgt cgaggacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca
     1861 tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagcttccag tccgtcctga
     1921 gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg
     1981 ggatcactct cggcatggac gagctgtaca acgtggatgg cggtagcggt ggcaccggca
     2041 gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg
     2101 taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc
     2161 tgaccctgaa gctgatctgc accaccggca agctgcccgt gccctggccc accctcgtga
     2221 ccaccctcgg ctacggcctg aagtgcttcg cccgctaccc cgaccacatg aagcagcacg
     2281 acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg
     2341 acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc
     2401 gcatcgagct gaagggcatc ggcttcaagg aggacggcaa catcctgggg cacaagctgg
     2461 agtacaacgg caccggtttc tgttttgaag ccggggcgga tgaagataca cacttccgcg
     2521 cgaccagcct ggaaactctg cgcaacatgg tggcggcagg tagcgggatc actttactgc
     2581 cagcgctggc tgtgccgccg gagcgcaaac gcgatggggt tgtttatctg ccgtgcatta
     2641 agccggaacc acgccgcact attggtctgg tttatcgtcc tggctcaccg ctgcgcagcc
     2701 gctatgagca gctggcagag gccatccgcg caagaatgga tggccatttc gataaagttt
     2761 taaaacaggc ggtttaaaag cttgcctcga gcagcgctgc tcgagagatc tacgggtggc
     2821 atccctgtga cccctcccca gtgcctctcc tggccctgga agttgccact ccagtgccca
     2881 ccagccttgt cctaataaaa ttaagttgca tcattttgtc tgactaggtg tccttctata
     2941 atattatggg gtggaggggg gtggtatgga gcaaggggca agttgggaag acaacctgta
     3001 gggcctgcgg ggtctattgg gaaccaagct ggagtgcagt ggcacaatct tggctcactg
     3061 caatctccgc ctcctgggtt caagcgattc tcctgcctca gcctcccgag ttgttgggat
     3121 tccaggcatg catgaccagg ctcagctaat ttttgttttt ttggtagaga cggggtttca
     3181 ccatattggc caggctggtc tccaactcct aatctcaggt gatctaccca ccttggcctc
     3241 ccaaattgct gggattacag gcgtgaacca ctgctccctt ccctgtcctt ctgattttgt
     3301 aggtaaccac gtgcggaccg agcggccgca ggaaccccta gtgatggagt tggccactcc
     3361 ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc gacgcccggg
     3421 ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc tgcctgcagg ggcgcctgat
     3481 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatacgtc aaagcaacca
     3541 tagtacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     3601 accgctacac ttgccagcgc cttagcgccc gctcctttcg ctttcttccc ttcctttctc
     3661 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     3721 tttagtgctt tacggcacct cgaccccaaa aaacttgatt tgggtgatgg ttcacgtagt
     3781 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     3841 agtggactct tgttccaaac tggaacaaca ctcaactcta tctcgggcta ttcttttgat
     3901 ttataaggga ttttgccgat ttcggtctat tggttaaaaa atgagctgat ttaacaaaaa
     3961 tttaacgcga attttaacaa aatattaacg tttacaattt tatggtgcac tctcagtaca
     4021 atctgctctg atgccgcata gttaagccag ccccgacacc cgccaacacc cgctgacgcg
     4081 ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg
     4141 agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc
     4201 gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt
     4261 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     4321 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     4381 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     4441 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     4501 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     4561 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     4621 ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     4681 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     4741 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     4801 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     4861 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     4921 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     4981 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     5041 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     5101 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     5161 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     5221 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     5281 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     5341 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     5401 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     5461 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     5521 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     5581 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     5641 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     5701 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     5761 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     5821 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     5881 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     5941 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     6001 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     6061 cacatgt
//
