LOCUS       sequence-491401-        6985 bp    DNA     circular SYN 03-NOV-2025
DEFINITION  3' AAVLINK plasmid for SYNGAP1 expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6985)
  AUTHORS   Zhonghua Lu Lab
  TITLE     AAVLINK: A Potent DNA-Recombination Method for Large Cargo Delivery 
            in Gene Therapy
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6985)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 3, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pAAVLINKv1-pCALM1-Cre-3'-SYNGAP1-FLAG
FEATURES             Location/Qualifiers
     source          1..6985
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     CDS             277..1308
                     /codon_start=1
                     /product="site-specific recombinase"
                     /label=Cre
                     /note="Cre is a site-specific recombinase from 
                     bacteriophage P1. Recombination occurs at loxP sequences."
                     /translation="MSNLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFSEHTWKML
                     LSVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGL
                     PRPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQDIRNLAF
                     LGIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSLGVTKLVE
                     RWISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEATHRLIYGAKDDSGQ
                     RYLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRNLDSETGAMVRLLE
                     DGD"
     primer_bind     1464..1483
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             2649..2678
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             3705..3770
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(3835..3855)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   4248..4388
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4248..4377
                     /label=AAV2 ITR
     rep_origin      4463..4918
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4550..4569)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4760..4781
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4935..4954)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5054..5076
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5114..5132)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5200..5304
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5305..5373,5374..6165)
                     /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(5523..5542)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6336..6924
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6825..6844
                     /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 gcgtcgctct gcgcgaggcg gctccgccgc ggcggaggga
      181 tacggcgcac catatatata tcgcggggcg cagactcgcg ctccggcagt ggtgctggga
      241 gtgtcgtgga cgccgtgccg ttactcgtac gccaccatgt ccaatttact gaccgtacac
      301 caaaatttgc ctgcattacc ggtcgatgca acgagtgatg aggttcgcaa gaacctgatg
      361 gacatgttca gggatcgcca ggcgttttct gagcatacct ggaaaatgct tctgtccgtt
      421 tgccggtcgt gggcggcatg gtgcaagttg aataaccgga aatggtttcc cgcagaacct
      481 gaagatgttc gcgattatct tctatatctt caggcgcgcg gtctggcagt aaaaaccatc
      541 cagcaacatt tgggccagct aaacatgctt catcgtcggt ccgggctgcc acgaccaagt
      601 gacagcaatg ctgtttcact ggttatgcgg cggatccgaa aagaaaacgt tgatgccggt
      661 gaacgtgcaa aacaggctct agcgttcgaa cgcactgatt tcgaccaggt tcgttcactc
      721 atggaaaata gcgatcgctg ccaggatata cgtaatctgg catttctggg gattgcttat
      781 aacaccctgt tacgtatagc cgaaattgcc aggatcaggg ttaaagatat ctcacgtact
      841 gacggtggga gaatgttaat ccatattggc agaacgaaaa cgctggttag caccgcaggt
      901 gtagagaagg cacttagcct gggggtaact aaactggtcg agcgatggat ttccgtctct
      961 ggtgtagctg atgatccgaa taactacctg ttttgccggg tcagaaaaaa tggtgttgcc
     1021 gcgccatctg ccaccagcca gctatcaact cgcgccctgg aagggatttt tgaagcaact
     1081 catcgattga tttacggcgc taaggatgac tctggtcaga gatacctggc ctggtctgga
     1141 cacagtgccc gtgtcggagc cgcgcgagat atggcccgcg ctggagtttc aataccggag
     1201 atcatgcaag ctggtggctg gaccaatgta aatattgtaa tgaactatat ccgtaacctg
     1261 gatagtgaaa caggggcaat ggtgcgcctg ctggaagatg gcgattaggt aagtatcaag
     1321 gttacaagac aggtttaagg agaccaataa ctagtataac ttcgtatagg atactttata
     1381 gcaatttatg aaactgggct tgtcgagaca gagaagactc ttgcgtttct gataggcacc
     1441 tattggtctt actgacatcc actttgcctt tctctccaca ggtggtcaac tcccactgcg
     1501 tgttcccgag ggagctgaag gaggtgtttg cttcatggcg gctgcgctgc gcagagcgag
     1561 gccgggagga catcgcagac aggcttatca gcgcctcact cttcctgcgc ttcctctgcc
     1621 cagcgattat gtcgcccagt ctctttgggc ttatgcagga gtacccagat gagcagacct
     1681 cacgaaccct caccctcatt gccaaggtca tccagaacct ggccaacttt tccaagttta
     1741 cctcaaagga ggactttctg ggcttcatga atgagtttct ggagctggaa tggggttcca
     1801 tgcagcagtt tttgtatgag atctccaatc tggacacgct aaccaacagc agtagctttg
     1861 agggttacat cgacttgggc cgagagctct ccacactgca tgccctactc tgggaggtgc
     1921 tgccccagct cagcaaggaa gccctcctga agctgggtcc actgccccgg ctcctcaacg
     1981 acatcagcac agctctgagg aaccccaaca tccaaaggca gccaagccgc cagagtgagc
     2041 ggccccggcc tcagcctgtg gtactgcggg ggccatcggc tgagatgcag ggctacatga
     2101 tgcgggacct caacagctct atggacatgg ctcgcctccc ctccccaacc aaggaaaagc
     2161 cacccccacc accgcctggt ggtggtaaag acctgttcta tgtaagccgt ccacccctgg
     2221 cccgttcctc accagcatac tgcacgagca gctcggacat cacagagcca gagcagaaga
     2281 tgctgagtgt caacaagagt gtgtccatgc tggacttaca gggtgatggg cctggtggcc
     2341 gcctcaacag cagcagtgtt tcgaacctgg cggccgtagg ggacctgctg cactcaagcc
     2401 aggcctcgct gacagcagcc ttggggctac ggcctgcgcc tgccggacgc ctctcccagg
     2461 ggagtggctc atccatcacg gcggctggca tgcgcctcag ccagatgggt gtcaccacag
     2521 acggtgtccc tgcccagcaa ctgcgaatcc ccctctcctt ccagaaccct ctcttccaca
     2581 tggctgctga tgggccaggt cccccaggcg gccatggagg gggcggtggc catggcccac
     2641 cttcctccca tcaccaccac caccaccatc accaccaccg aggtggagag ccccctgggg
     2701 acacctttgc cccattccat ggctatagca agagtgagga cctctcttcc ggggtcccca
     2761 agccccctgc tgcctccatc cttcatagcc acagctacag tgatgagttt ggaccctctg
     2821 gcactgactt cacccgtcgg cagctttcac tccaggacaa cctgcagcac atgctgtccc
     2881 ctccccagat caccattggt ccccagaggc cagccccctc agggcctgga ggtgggagcg
     2941 gtgggggcag cggtgggggt ggcgggggcc agccgcctcc attgcagagg ggcaagtctc
     3001 agcagttgac agtcagcgca gcccagaaac cccggccatc cagcgggaat ctattgcagt
     3061 ccccagagcc aagttatggc cccgcccgtc cacggcaaca gagcctcagc aaggagggca
     3121 gcattggggg cagcgggggc agcggtggcg gagggggtgg ggggctgaag ccctccatca
     3181 ccaagcagca ttctcagaca ccatccacat tgaaccccac aatgccagcc tctgagcgga
     3241 cagtggcctg ggtctccaac atgcctcacc tgtcggctga catcgagagt gcccacatcg
     3301 agcgggaaga gtacaagctc aaggagtact caaaatcgat ggatgagagc cggctggata
     3361 gggagtacga ggaggagatt cactcactga aagagcggct gcacatgtcc aaccggaagc
     3421 tggaagagta tgagcggagg ctgctgtccc aggaagaaca aaccagcaaa atcctgatgc
     3481 agtatcaggc ccgactggag cagagtgaga agaggctaag gcagcagcag gcagagaagg
     3541 attcccagat caagagcatc attggcaggc tgatgctggt ggaggaggag ctgcgccggg
     3601 accaccccgc catggctgag ccgctgccag aacccaagaa gaggctgctc gacgctcaga
     3661 gaggcagctt ccccccttgg gtccaacaaa cccgcgtgtg tacagactac aaagaccatg
     3721 acggtgatta taaagatcat gacatcgact acaaggatga cgatgacaag taactcgaga
     3781 taatcaacct ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc
     3841 tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta ttgcttcccg
     3901 tatggctttc attttctcct ccttgtataa atcctggtta gttcttgcca cggcggaact
     3961 catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc
     4021 cgtggtgttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca tctagcttta
     4081 tttgtgaaat ttgtgatgct attgctttat ttgtaaccat tataagctgc aataaacaag
     4141 ttaacaacaa caattgcatt cattttatgt ttcaggttca gggggagatg tgggaggttt
     4201 tttaaagatc tggagaccgg tgtggcacgt gcggaccgag cggccgcagg aacccctagt
     4261 gatggagttg gccactccct ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa
     4321 ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg
     4381 cctgcagggg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     4441 catacgtcaa agcaaccata gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     4501 gtggttacgc gcagcgtgac cgctacactt gccagcgcct tagcgcccgc tcctttcgct
     4561 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     4621 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgatttg
     4681 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     4741 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caactctatc
     4801 tcgggctatt cttttgattt ataagggatt ttgccgattt cggtctattg gttaaaaaat
     4861 gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt tacaatttta
     4921 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg
     4981 ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa
     5041 gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc
     5101 gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg
     5161 gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     5221 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     5281 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     5341 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     5401 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     5461 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     5521 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     5581 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     5641 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     5701 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     5761 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     5821 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     5881 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     5941 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     6001 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     6061 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     6121 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     6181 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     6241 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     6301 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     6361 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     6421 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     6481 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     6541 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     6601 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     6661 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     6721 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     6781 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     6841 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     6901 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     6961 ttttgctggc cttttgctca catgt
//
