LOCUS       Exported                6439 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  AAV expression of Cre recombinase-activated jRGECO1a, a red 
            fluorescent calcium sensor protein, from the CAG promoter.
ACCESSION   .
VERSION     .
KEYWORDS    pAAV.CAG.Flex.NES-jRGECO1a.WPRE.SV40
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6439)
  AUTHORS   Dana H, Mohar B, Sun Y, Narayan S, Gordus A, Hasseman JP, Tsegaye G,
            Holt GT, Hu A, Walpita D, Patel R, Macklin JJ, Bargmann CI, Ahrens 
            MB, Schreiter ER, Jayaraman V, Looger LL, Svoboda K, Kim DS
  TITLE     Sensitive red protein calcium indicators for imaging neural 
            activity.
  JOURNAL   Elife. 2016 Mar 24;5. pii: e12727. doi: 10.7554/eLife.12727.
  PUBMED    27011354
REFERENCE   2  (bases 1 to 6439)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6439
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     repeat_region   1..130
                     /label=AAV2 ITR
     promoter        441..718
                     /label=chicken beta-actin promoter
     primer_bind     997..1019
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1081..1100
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    complement(1135..1168)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites (Lee and 
                     Saito, 1988)."
     protein_bind    complement(1231..1264)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     primer_bind     complement(1742..1761)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             complement(2457..2513)
                     /codon_start=1
                     /product="calmodulin-binding peptide from avian smooth 
                     muscle myosin light chain kinase"
                     /label=calmodulin-binding peptide
                     /translation="RRKWNKAGHAVRAIGRLSS"
     CDS             complement(2541..2564)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             complement(2568..2600)
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             complement(2604..2621)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     protein_bind    2704..2737
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites (Lee and 
                     Saito, 1988)."
     protein_bind    2808..2841
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    2883..3471
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2936..2956)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3354..3365)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3514..3635
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3525..3544)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3579..3598
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   3797..3926
                     /label=AAV2 ITR
     primer_bind     complement(3992..4009)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4163..4751)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4243..4262)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4922..5782)
                     /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     5545..5564
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5783..5887)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5913..6368)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6050..6071)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6262..6281
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct tgtagttaat gattaacccg ccatgctact tatctacgta gccatgctct
      181 aggaagagta ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
      241 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
      301 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
      361 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt
      421 agtcatcgct attaccatgg tcgaggtgag ccccacgttc tgcttcactc tccccatctc
      481 ccccccctcc ccacccccaa ttttgtattt atttattttt taattatttt gtgcagcgat
      541 gggggcgggg gggggggggg ggcgcgcgcc aggcggggcg gggcggggcg aggggcgggg
      601 cggggcgagg cggagaggtg cggcggcagc caatcagagc ggcgcgctcc gaaagtttcc
      661 ttttatggcg aggcggcggc ggcggcggcc ctataaaaag cgaagcgcgc ggcgggcggg
      721 agtcgctgcg cgctgccttc gccccgtgcc ccgctccgcc gccgcctcgc gccgcccgcc
      781 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc
      841 gggctgtaat tagcgcttgg tttaatgacg gcttgtttct tttctgtggc tgcgtgaaag
      901 ccttgagggg ctccgggagg gccctttgtg cggggggagc ggctcggggc tgtccgcggg
      961 gggacggctg ccttcggggg ggacggggca gggcggggtt cggcttctgg cgtgtgaccg
     1021 gcggctctag agcctctgct aaccatgttc atgccttctt ctttttccta cagctcctgg
     1081 gcaacgtgct ggttattgtg ctgtctcatc attttggcaa agaattggat ccccataact
     1141 tcgtataaag tatcctatac gaagttatat caaaatagga agaccaatgc ttcaccatcg
     1201 acccgaattg ccaagcatca ccatcgaccc ataacttcgt ataatgtatg ctatacgaag
     1261 ttatactaga gctttcactt cgctgtcatc atttgtacaa actcttcgta gtttacctga
     1321 ccatccccat cgatgtctgc taccctgatc atttcatcaa cctcctcatc tgttaacttc
     1381 tctccaaggt ctgtcatcac gtggcgaagc tctgctgcgc cgatgtagcc attgccgtcc
     1441 ttatcaaaca cgcggaacgc ttcgcgaatt tcctcttcac tgtctgtgtc attcattttt
     1501 cttgccatca tcgtcaggaa ctcagggaag tcgaatgtgc cgtcaccgtc ggcatctact
     1561 tcattgatca tgtcctgcag ctctgcttct gtggggttct gccccagcga tcggaacacc
     1621 gtcccaagct ccttggttgt tatcgtccca tccccgtcct tgtcaaatag ggagaaagcc
     1681 tctttaaatt ctgcgatctg ctcttcagtc aggtcgtcac gcgtagcctc ccagcccatg
     1741 gtcttcttct gcattacggg gccgtcgggg gggaagttgg tgccgcgcag cttcaccttg
     1801 tagatgaata cgccgtcctg cagggaggag tcctggttca cgtgaataat gccgccgtcc
     1861 tcgaagttca tcacgcgctc ccacctgaag ccctcgggga aggacagctt gaagtagtcg
     1921 gggatgtcgg ctgggtgctt aatgtaggcc ttggagccgt acatgaactg aggggacagg
     1981 atgtcccagg cgaagggcag ggggccaccc ttggtcacct tcagcttagc ggtctgaaag
     2041 gcctcgtagg ggcggccctc gccctcgccc tcgatctcga actcgtggcc gttcacggag
     2101 ccctccatgt gcaccttgaa gcgcatgaac tccttgatga tggccatgtt atcctcctcg
     2161 cccttgctca ccagactccc gcctgtacct cccttgtaca gctcgtccat gccgccggtg
     2221 gagtggcggc cctcggcgcg ttcgcactgt tccacgatgg tgtagtcctc gttgtgggac
     2281 acgatgtcca acttgatgtc cacgatgtag gcgccgggca gctgcacggg cttcttggcc
     2341 ttgtaggtgg tcttgacctc ggcggcgtag tggccgccgt ccttcagcct cagccccttc
     2401 ttgatctcgc tcttcagggc gccgtcctcg gggtacatcc gctcggaaac cacgggtgag
     2461 ctcagccgac ctatagctct gactgcgtga cctgccttat tccacttacg acgtgatgag
     2521 tcgaccattg ttgcgagatc cttatcgtca tcgtcgtaca gatcccgacc catttgctgt
     2581 ccaccagtca tgctagccat accatgatga tgatgatgat gagaacctcc tccagtcttg
     2641 ttaatatcaa gtccagccaa cttaagagca agctcgttct gcagcatggt ggcggatcct
     2701 agtataactt cgtataggat actttatacg aagttatcat tgggattctt cctattttga
     2761 tccaagcatc accatcgacc ctctagtcca gatctcacca tcgacccata acttcgtata
     2821 gcatacatta tacgaagtta tgtccctcga agaggttcga attcgatatc aagcttatcg
     2881 ataatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt aactatgttg
     2941 ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct attgcttccc
     3001 gtatggcttt cattttctcc tccttgtata aatcctggtt gctgtctctt tatgaggagt
     3061 tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca
     3121 ctggttgggg cattgccacc acctgtcagc tcctttccgg gactttcgct ttccccctcc
     3181 ctattgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca ggggctcggc
     3241 tgttgggcac tgacaattcc gtggtgttgt cggggaaatc atcgtccttt ccttggctgc
     3301 tcgcctgtgt tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc
     3361 tcaatccagc ggaccttcct tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc
     3421 ttcgccttcg ccctcagacg agtcggatct ccctttgggc cgcctccccg catcgatacc
     3481 gtcgacccgg gcggccgctt cgagcagaca tgataagata cattgatgag tttggacaaa
     3541 ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt
     3601 tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta
     3661 tgtttcaggt tcagggggag atgtgggagg ttttttaaag caagtaaaac ctctacaaat
     3721 gtggtaaaat cgataaggat cttcctagag catggctacg tagataagta gcatggcggg
     3781 ttaatcatta actacaagga acccctagtg atggagttgg ccactccctc tctgcgcgct
     3841 cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg
     3901 gcctcagtga gcgagcgagc gcgcagctgc attaatgaat cggccaacgc gcggggagag
     3961 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     4021 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     4081 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     4141 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     4201 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     4261 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     4321 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     4381 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     4441 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     4501 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     4561 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     4621 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     4681 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     4741 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     4801 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     4861 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     4921 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     4981 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     5041 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     5101 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     5161 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     5221 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     5281 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     5341 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     5401 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     5461 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     5521 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     5581 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     5641 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     5701 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     5761 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     5821 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     5881 ttccgcgcac atttccccga aaagtgccac ctaaattgta agcgttaata ttttgttaaa
     5941 attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa
     6001 aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa
     6061 caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca
     6121 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg
     6181 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc
     6241 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc
     6301 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca
     6361 gggcgcgtcc cattcgccat tcaggctgcg caactgttgg gaagggcgat cggtgcgggc
     6421 ctcttcgcta ttacgccag
//
