LOCUS       sequence-471024-        6911 bp    DNA     circular SYN 17-DEC-2025
DEFINITION  to express the optogenetic tool 2pabPAC, which increases cAMP levels
            when stimulated by blue light, in astrocytes.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6911)
  AUTHORS   Vittani M, Herlo R, Wang X, Christensen MDB, Vo CT, Mishima T, Kusk 
            P, Konno A, Hirai H, Tsuboi T, Kitaguchi T, Kjaerby C, Asiminas A, 
            Yokoyama T, Sakamoto M, Nedergaard M, Hirase H
  TITLE     Cerebral blood flow is modulated by astrocytic cAMP elevation 
            independently of IP(3)R2-mediated Ca(2+) signaling in mice.
  JOURNAL   Proc Natl Acad Sci U S A. 2025 Jul 8;122(27):e2422069122. doi: 
            10.1073/pnas.2422069122. Epub 2025 Jul 1.
  PUBMED    40591593
REFERENCE   2  (bases 1 to 6911)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 17, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Proc Natl 
            Acad Sci U S A."; date: "2025-07-8"; volume: "122(27):e2422069122. 
            doi"; pages: " 10.1073/pnas.2422069122. Epub 2025 Jul 1"
COMMENT     Sequence Label: pAAV-mGFAP(ABC1D)-2pabPAC
FEATURES             Location/Qualifiers
     source          1..6911
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   192..332
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   192..321
                     /label=AAV2 ITR
     rep_origin      407..862
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(494..513)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     704..725
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(879..898)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     998..1020
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1058..1076)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1144..1248
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1249..1317,1318..2109)
                     /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(1467..1486)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2280..2868
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2769..2788
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   3105..3234
                     /label=AAV2 ITR
     CDS             5501..6193
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet
                     /translation="VSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSFPEGFKWERVMNFEDGGA
                     VTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIKM
                     ALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRHS
                     TGGMDELYK"
     primer_bind     complement(5631..5649)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     5899..5918
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    6228..6816
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(6281..6301)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6699..6710)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    join(6834..6911,1..147)
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
ORIGIN
        1 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta
       61 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc
      121 atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagatctacc acgtgcggac
      181 cgagcggccg caggaacccc tagtgatgga gttggccact ccctctctgc gcgctcgctc
      241 gctcactgag gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc
      301 agtgagcgag cgagcgcgca gctgcctgca ggggcgcctg atgcggtatt ttctccttac
      361 gcatctgtgc ggtatttcac accgcatacg tcaaagcaac catagtacgc gccctgtagc
      421 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
      481 gccttagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
      541 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
      601 ctcgacccca aaaaacttga tttgggtgat ggttcacgta gtgggccatc gccctgatag
      661 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
      721 actggaacaa cactcaactc tatctcgggc tattcttttg atttataagg gattttgccg
      781 atttcggtct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac
      841 aaaatattaa cgtttacaat tttatggtgc actctcagta caatctgctc tgatgccgca
      901 tagttaagcc agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg
      961 ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg
     1021 ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg cctattttta
     1081 taggttaatg tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat
     1141 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     1201 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa
     1261 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac
     1321 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac
     1381 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt
     1441 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc
     1501 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca
     1561 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc
     1621 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag
     1681 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa
     1741 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg
     1801 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa
     1861 ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg
     1921 gctggctggt ttattgctga taaatctgga gccggtgagc gtggaagccg cggtatcatt
     1981 gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt
     2041 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag
     2101 cattggtaac tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat
     2161 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     2221 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     2281 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     2341 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     2401 agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc
     2461 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     2521 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     2581 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     2641 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     2701 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     2761 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     2821 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     2881 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
     2941 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc
     3001 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcccaata
     3061 cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctgcgc gctcgctcgc
     3121 tcactgaggc cgcccgggca aagcccgggc gtcgggcgac ctttggtcgc ccggcctcag
     3181 tgagcgagcg agcgcgcaga gagggagtgg ccaactccat cactaggggt tccttgtagt
     3241 taatgattaa cccgccatgc tacttatcta ctcgagaccg ggcgcgccgg gggaggctgc
     3301 tggtgaatat taaccaaggt caccccagtt atcggaggag caaacagggg ctaagtccac
     3361 acgcgtggta ccgtctgtct gcacatttcg tagagcgagt gttccgatac tctaatctcc
     3421 ctaggcaagg ttcatatttg tgtaggttac ttattctcct tttgttgact aagtcaataa
     3481 tcagaatcag caggtttgga gtcagcttgg cagggatcag cagcctgggt tggaaggagg
     3541 gggtataaaa gccccttcac caggagaagc cgtcacacag atccacaagc tcctggtcga
     3601 cactagtacc ggtgccacca tgaagtgggt aacctttctc ctcctcctct tcgtctccgg
     3661 ctctgctttt tccaggggtg tgtttcgccg agaagcacac aagagtgaga tcgcccatcg
     3721 gtataatgat ttgggagaac aacatttcaa aggcctagtc ctgattgcct tttcccagta
     3781 tctccagaaa tgctcatacg atgagcatgc caaattagtg caggaagtaa cagactttgc
     3841 aaagacgtgt gttgccgatg agtctgccgc caactgtgac aaatcccttc acactctttt
     3901 tggagataag ttgtgtgcca ttccaaacct ccgtgaaaac tatggtgaac tggctgactg
     3961 ctgtacaaaa caagagcccg aaagaaacga atgtttcctg caacacaaag atgacaaccc
     4021 cagcctgcca ccatttgaaa ggccagaggc tgaggccatg tgcacctcct ttaaggaaaa
     4081 cccaaccacc tttatgggac actatttgca tgaagttgcc agaagacatc cttatttcta
     4141 tgccccagaa cttctttact atgctgagca gtacaatgag attctgaccc agtgttgtgc
     4201 agaggctgac aaggaaagct gcctgacccc gaagcttgat ggtgtgaagg agaaagcatt
     4261 ggtctcatct gtccgtcaga gaatgaagtg ctccagtatg cagaagtttg gagagagagc
     4321 ttttaaagca tgggcagtag ctcgtctgag ccagacattc cccaatgctg actttgcaga
     4381 aatcaccaaa ttggcaacag acctgaccaa agtcaacaag gagtgctgcc atggtgacct
     4441 gctggaatgc gcagatgaca gggcggaact tgccaagtac atgtgtgaaa accaggcgac
     4501 tatctccagc aaactgcaga cttgctgcga taaaccactg ttgaagaaag cccactgtct
     4561 tagtgaggtg gagcatgaca ccatgcctgc tgatctgcct gccattgctg ctgattttgt
     4621 tgaggaccag gaagtgtgca agaactatgc tgaggccaag gatgtcttcc tgggcacgtt
     4681 cttgtatgaa tattcaagaa gacaccctga ttactctgta tccctgttgc tgagacttgc
     4741 taagaaatat gaagccactc tggaaaagtg ctgcgctgaa gccaatcctc ccgcatgcta
     4801 cggcacagtg cttgctgaat ttcagcctct tgtagaagag cctaagaact tggtcaaaac
     4861 caactgtgat ctttacgaga agcttggaga atatggattc caaaatgcca ttctagttcg
     4921 ctacacccag aaagcacctc aggtgtcaac cccaactctc gtggaggctg caagaaacct
     4981 aggaagagtg ggcaccaagt gttgtacact tcctgaagat cagagactgc cttgtgtgga
     5041 agactatctg tctgcaatcc tgaaccgtgt gtgtctgctg catgagaaga ccccagtgag
     5101 tgagcatgtt accaagtgct gtagtggatc cctggtggaa aggcggccat gcttctctgc
     5161 tctgacagtt gatgaaacat atgtccccaa agagtttaaa gctgagacct tcaccttcca
     5221 ctctgatatc tgcacacttc cagagaagga gaagcagatt aagaaacaaa cggctcttgc
     5281 tgagctggtg aagcacaagc ccaaggctac agcggagcaa ctgaagactg tcatggatga
     5341 ctttgcacag ttcctggata catgttgcaa ggctgctgac aaggacacct gcttctcgac
     5401 tgagggtcca aaccttgtca ctagatgcaa agacgcctta gcccccgggg gtggaggcgg
     5461 ttcaggcgga ggtggctctg gcggtggcgg atcagtcgac gtgagcaagg gcgaggcagt
     5521 gatcaaggag ttcatgcggt tcaaggtgca catggagggc tccatgaacg gccacgagtt
     5581 cgagatcgag ggcgagggcg agggccgccc ctacgagggc acccagaccg ccaagctgaa
     5641 ggtgaccaag ggtggccccc tgcccttctc ctgggacatc ctgtcccctc agttcatgta
     5701 cggctccagg gccttcacca agcaccccgc cgacatcccc gactactata agcagtcctt
     5761 ccccgagggc ttcaagtggg agcgcgtgat gaacttcgag gacggcggcg ccgtgaccgt
     5821 gacccaggac acctccctgg aggacggcac cctgatctac aaggtgaagc tccgcggcac
     5881 caacttccct cctgacggcc ccgtaatgca gaagaagaca atgggctggg aagcgtccac
     5941 cgagcggttg taccccgagg acggcgtgct gaagggcgac attaagatgg ccctgcgcct
     6001 gaaggacggt ggccgctacc tggcggactt caagaccacc tacaaggcca agaagcccgt
     6061 gcagatgccc ggcgcctaca acgtcgaccg caagttggac atcacctccc acaacgagga
     6121 ctacaccgtg gtggaacagt acgaacgctc cgagggccgc cactccaccg gcggcatgga
     6181 cgagctgtac aagtagcggc cgcgaattcg atatcaagct tatcgataat caacctctgg
     6241 attacaaaat ttgtgaaaga ttgactggta ttcttaacta tgttgctcct tttacgctat
     6301 gtggatacgc tgctttaatg cctttgtatc atgctattgc ttcccgtatg gctttcattt
     6361 tctcctcctt gtataaatcc tggttgctgt ctctttatga ggagttgtgg cccgttgtca
     6421 ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt tggggcattg
     6481 ccaccacctg tcagctcctt tccgggactt tcgctttccc cctccctatt gccacggcgg
     6541 aactcatcgc cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca
     6601 attccgtggt gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc tgtgttgcca
     6661 cctggattct gcgcgggacg tccttctgct acgtcccttc ggccctcaat ccagcggacc
     6721 ttccttcccg cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc
     6781 agacgagtcg gatctccctt tgggccgcct ccccgctctc gagatacgta cgactgtgcc
     6841 ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg
     6901 tgccactccc a
//
