LOCUS       SK0844(pAAV-GfaA        6805 bp    DNA     circular SYN 22-OCT-2025
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 6805)
  AUTHORS   Institute for Basic Science
  TITLE     Direct Submission
  JOURNAL   Exported Oct 22, 2025 from SnapGene 8.1.1
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6805
                     /mol_type="other DNA"
                     /organism="recombinant plasmid"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
                     /note="color: #ffe4c4"
     primer_bind     150..180
                     /label=MluI-gfaABC1D-JSKF
                     /note="color: black; sequence: 
                     atcgACGCGTAACATATCCTGGTGTGGAGTAGGGG"
     misc_feature    156..836
                     /label=GfaABC1D
                     /note="color: #ff99cc"
     primer_bind     complement(825..842)
                     /label=gfABC1D-XbaI-JSKR
                     /note="color: black; sequence: atcgTCTAGAGCGAGCAGCGGA"
     CDS             join(874..876,877..879,880..1590)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /note="This feature has 3 segments:
                        1:  874 ..  876 / #05fd14
                        2:  877 ..  879 / #05fd14 / 1a
                        3:  880 .. 1590 / #05fd14"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     1588..1608
                     /label=FLAG-CRY2(EA9,NheI)-F
                     /note="color: black; sequence: 
                     GACTGCTAGCGCCACCATGGGAGACTACAAGGATGACGACGATAAGCCACCGGTCATGA
                     AGATG; added: 2022-02-13"
     CDS             1600..3093
                     /codon_start=1
                     /label=CRY2(E281A)
                     /note="color: #33cccc"
                     /translation="MKMDKKTIVWFRRDLRIEDNPALAAAAHEGSVFPVFIWCPEEEGQ
                     FYPGRASRWWMKQSLAHLSQSLKALGSDLTLIKTHNTISAILDCIRVTGATKVVFNHLY
                     DPVSLVRDHTVKEKLVERGISVQSYNGDLLYEPWEIYCEKGKPFTSFNSYWKKCLDMSI
                     ESVMLPPPWRLMPITAAAEAIWACSIEELGLENEAEKPSNALLTRAWSPGWSNADKLLN
                     EFIEKQLIDYAKNSKKVVGNSTSLLSPYLHFGEISVRHVFQCARMKQIIWARDKNSEGA
                     ESADLFLRGIGLREYSRYICFNFPFTHEQSLLSHLRFFPWDADVDKFKAWRQGRTGYPL
                     VDAGMRELWATGWMHNRIRVIVSSFAVKFLLLPWKWGMKYFWDTLLDADLECDILGWQY
                     ISGSIPDGHELDRLDNPALQGAKYDPEGEYIRQWLPELARLPTEWIHHPWDAPLTVLKA
                     SGVELGTNYAKPIVDIDTARELLAKAISRTREAQIMIGAA"
     primer_bind     complement(2417..2448)
                     /label=STIM1(E281A)-R
                     /note="color: black; sequence: 
                     GCTTTCGGCGCCTTCGCTGTTTTTATCGCGAG"
     primer_bind     2434..2463
                     /label=PHR(E281A)-F
                     /note="color: black; sequence: 
                     GAAGGCGCCGAAAGCGCCGATCTGTTCCTG"
     CDS             3094..3120
                     /codon_start=1
                     /product="CRY2clust"
                     /label=9 amino acids
                     /note="color: #ffffff"
                     /translation="ARDPPDLDN"
     CDS             3199..3597
                     /codon_start=1
                     /label=STIM1(318-450)
                     /note="color: #ff0000"
                     /translation="EEELEQVREALRKAEKELESHSSWYAPEALQKWLQLTHEVEVQYY
                     NIKKQNAEKQLLVAKEGAEKIKKKRNTLFGTFHVAHSSSLDDVDHKILTAKQALSEVTA
                     ALRERLHRWQQIEILCGFQIVNNPGIHSL"
     primer_bind     3598..3640
                     /label=BamHI-XhoI-CW3SL-JGSF
                     /note="color: black; sequence: 
                     TGGATGGGCAGTTAGGGATCCctcgagataatcaacctctggattacaaaatttg"
     misc_feature    3615..4041
                     /label=W3SL
                     /note="Optimization of AAV expression cassettes to improve 
                     packaging capacity and transgene expression in neurons
                     WPRE3 + Late SV40
                     "
                     /note="color: #ffffff"
     primer_bind     complement(3668..3688)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
                     /note="color: black; sequence: CATAGCGTAAAAGGAGCAACA"
     primer_bind     complement(4030..4071)
                     /label=CW3SL-RsrII-JGSR
                     /note="color: black; sequence: 
                     tcctgcggccgctcggtccgtcctgcggccgctttaaaaaac"
     repeat_region   4068..4208
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
                     /note="color: #ffe4c4"
     rep_origin      4283..4738
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     promoter        5020..5124
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: RIGHT"
     CDS             join(5125..5193,5194..5985)
                     /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: 5125 .. 5193 / #ccffcc / signal sequence
                        2: 5194 .. 5985 / #ccffcc"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      6156..6744
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgtaacat atcctggtgt ggagtagggg
      181 acgctgctct gacagaggct cgggggcctg agctggctct gtgagctggg gaggaggcag
      241 acagccaggc cttgtctgca agcagacctg gcagcattgg gctggccgcc ccccagggcc
      301 tcctcttcat gcccagtgaa tgactcacct tggcacagac acaatgttcg gggtgggcac
      361 agtgcctgct tcccgccgca ccccagcccc cctcaaatgc cttccgagaa gcccattgag
      421 cagggggctt gcattgcacc ccagcctgac agcctggcat cttgggataa aagcagcaca
      481 gccccctagg ggctgccctt gctgtgtggc gccaccggcg gtggagaaca aggctctatt
      541 cagcctgtgc ccaggaaagg ggatcagggg atgcccaggc atggacagtg ggtggcaggg
      601 ggggagagga gggctgtctg cttcccagaa gtccaaggac acaaatgggt gaggggagag
      661 ctctccccat agctgggctg cggcccaacc ccaccccctc aggctatgcc agggggtgtt
      721 gccaggggca cccgggcatc gccagtctag cccactcctt cacaaagccc tcgcatccca
      781 ggagcgagca gagccagagc aggttggaga ggagacgcat cacctccgct gctcgctcta
      841 gagccgatcc ccgggtaccg ggctagcgcc accatggtga gcaagggcga ggagctgttc
      901 accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc
      961 gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc
     1021 accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg
     1081 cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg
     1141 cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc
     1201 cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc
     1261 gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac
     1321 aacgtctata tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc
     1381 cacaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc
     1441 ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc cgccctgagc
     1501 aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg
     1561 atcactctcg gcatggacga gctgtacaag ccaccggtca tgaagatgga caaaaagacc
     1621 atcgtctggt ttcggagaga tttgagaata gaagataatc ccgcgctcgc cgccgcggcc
     1681 cacgagggtt ccgtcttccc cgttttcatt tggtgtcctg aagaagaagg ccagttttat
     1741 cccggaaggg cctctaggtg gtggatgaag caaagtctgg cccatcttag ccagtcactg
     1801 aaagcactgg gcagtgatct taccctgatc aagacacaca ataccatctc tgccatcctc
     1861 gactgcatca gggtgaccgg cgcaacgaaa gtcgtgttta accacctgta cgatccagtt
     1921 agtctggtgc gcgaccacac tgtgaaggag aagctggtgg aacgggggat cagtgtgcag
     1981 agctacaacg gggaccttct gtacgagcca tgggagatct attgcgagaa agggaaaccg
     2041 ttcacctcct tcaacagtta ctggaagaaa tgtttggata tgtcaataga gtccgttatg
     2101 ttgccccctc cctggagact gatgccgatt actgctgctg cagaggccat ctgggcctgc
     2161 tccatcgagg aactcggtct ggaaaatgaa gcagaaaagc caagcaatgc acttctcact
     2221 agagcctgga gccccggctg gtctaatgcc gacaagctgc ttaacgagtt catcgaaaaa
     2281 caactgattg actacgcgaa gaactccaag aaagtggtag gtaactcaac tagcttgctc
     2341 tctccatatc tccattttgg cgagatttct gtccgccatg tatttcagtg cgctcggatg
     2401 aaacagatta tctgggctcg cgataaaaac agcgaaggcg ccgaaagcgc cgatctgttc
     2461 ctgcgaggga tcggacttcg ggaatactcc cggtatatat gtttcaactt tccattcaca
     2521 cacgagcaga gtctgttgtc ccacctcagg ttcttcccct gggacgccga tgtcgacaaa
     2581 ttcaaggcat ggagacaggg aaggacaggg tacccactcg tggatgctgg catgagagag
     2641 ctctgggcta caggctggat gcacaaccgc atccgggtaa tcgtgtcctc atttgctgtc
     2701 aagtttctgc tcctgccttg gaaatgggga atgaagtact tttgggatac ccttctcgac
     2761 gccgacttgg agtgtgacat tctgggatgg caatatatta gcgggtcaat tcctgacggc
     2821 catgagttgg acaggttgga caatccggcc ttgcagggag ctaagtatga tcccgaagga
     2881 gagtatattc gacagtggct ccccgagctg gcccgacttc ctacggagtg gattcaccat
     2941 ccttgggacg caccactgac agtgctcaag gcaagcgggg tggagctggg caccaattac
     3001 gctaagccta tagttgatat agatacagca cgcgagctgc tggctaaagc gatctctcgc
     3061 actcgggagg cgcagattat gatcggtgct gccgccaggg acccccccga cctcgacaac
     3121 gaattcagtg ggggaggcgg agggggtggt tcaggtggtg gtggtggttc gggtggtggc
     3181 ggagggggag gctccggaga ggaggagttg gagcaggttc gggaggcctt gaggaaagca
     3241 gagaaggagc tagaatctca cagctcatgg tatgctccag aggcccttca gaagtggctg
     3301 cagctgacac atgaggtgga ggtgcaatat tacaacatca agaagcaaaa tgctgagaag
     3361 cagctgctgg tggccaagga gggggctgag aagataaaaa agaagagaaa cacactcttt
     3421 ggcaccttcc acgtggccca cagctcttcc ctggatgatg tagatcataa aattctaaca
     3481 gctaagcaag cactgagcga ggtgacagca gcattgcggg agcgcctgca ccgctggcaa
     3541 cagatcgaga tcctctgtgg cttccagatt gtcaacaacc ctggcatcca ctcactgtag
     3601 ggatccctcg agataatcaa cctctggatt acaaaatttg tgaaagattg actggtattc
     3661 ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg
     3721 ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg ttagttcttg
     3781 ccacggcgga actcatcgcc gcctgccttg cccgctgctg gacaggggct cggctgttgg
     3841 gcactgacaa ttccgtggtg tttatttgtg aaatttgtga tgctattgct ttatttgtaa
     3901 ccatctagct ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc
     3961 tgcaataaac aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag
     4021 atgtgggagg ttttttaaag cggccgcagg acggaccgag cggccgcagg aacccctagt
     4081 gatggagttg gccactccct ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa
     4141 ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg
     4201 cctgcagggg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     4261 catacgtcaa agcaaccata gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     4321 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct
     4381 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     4441 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgatttg
     4501 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     4561 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc
     4621 tcgggctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
     4681 gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt tacaatttta
     4741 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg
     4801 ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa
     4861 gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc
     4921 gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg
     4981 gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     5041 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     5101 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     5161 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     5221 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     5281 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     5341 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     5401 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     5461 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     5521 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     5581 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     5641 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     5701 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     5761 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     5821 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     5881 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     5941 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     6001 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     6061 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     6121 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     6181 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     6241 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     6301 gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     6361 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     6421 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     6481 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     6541 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     6601 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     6661 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     6721 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     6781 ttttgctggc cttttgctca catgt
//
