LOCUS       sequence-491772-        6566 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  3' AAVLINK plasmid for GRIN2A expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6566)
  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  JOURNAL   Unpublished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 6566)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 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            "Unpublished"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'-GRIN2A-FLAG
FEATURES             Location/Qualifiers
     source          1..6566
                     /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"
     CDS             3286..3351
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(3416..3436)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   3829..3969
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3829..3958
                     /label=AAV2 ITR
     rep_origin      4044..4499
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4131..4150)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4341..4362
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4516..4535)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4635..4657
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4695..4713)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4781..4885
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4886..4954,4955..5746)
                     /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(5104..5123)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5917..6505
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6406..6425
                     /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 ggggcatcta cagctgcatt
     1501 catggagtgc acattgaaga aaagaagaag tctccagact tcaatctgac gggatcccag
     1561 agcaacatgt taaaactcct ccggtcagcc aaaaacattt ccagcatgtc caacatgaac
     1621 tcctcaagaa tggactcacc caaaagagct gctgacttca tccaaagagg ttccctcatc
     1681 atggacatgg tttcagataa ggggaatttg atgtactcag acaacaggtc ctttcagggg
     1741 aaagagagca tttttggaga caacatgaac gaactccaaa catttgtggc caaccggcag
     1801 aaggataacc tcaataacta tgtattccag ggacaacatc ctcttactct caatgagtcc
     1861 aaccctaaca cggtggaggt ggccgtgagc acagaatcca aagcgaactc tagaccccgg
     1921 cagctgtgga agaaatccgt ggattccata cgccaggatt cactatccca gaatccagtc
     1981 tcccagaggg atgaggcaac agcagagaat aggacccact ccctaaagag ccctaggtat
     2041 cttccagaag agatggccca ctctgacatt tcagaaacgt caaatcgggc cacgtgccac
     2101 agggaacctg acaacagtaa gaaccacaaa accaaggaca actttaaaag gtcagtggcc
     2161 tccaaatacc ccaaggactg tagtgaggtc gagcgcacct acctgaaaac caaatcaagc
     2221 tcccctagag acaagatcta cactatagat ggtgagaagg agcctggttt ccacttagat
     2281 ccaccccagt ttgttgaaaa tgtgaccctg cccgagaacg tggacttccc ggacccctac
     2341 caggatccca gtgaaaactt ccgcaagggg gactccacgc tgccaatgaa ccggaacccc
     2401 ttgcataatg aagaggggct ttccaacaac gaccagtata aactctactc caagcacttc
     2461 accttgaaag acaagggttc cccgcacagt gagaccagcg agcgataccg gcagaactcc
     2521 acgcactgca gaagctgcct ttccaacatg cccacctatt caggccactt caccatgagg
     2581 tcccccttca agtgcgatgc ctgcctgcgg atggggaacc tctatgacat cgatgaagac
     2641 cagatgcttc aggagacagg taacccagcc accggggagc aggtctacca gcaggactgg
     2701 gcacagaaca atgcccttca attacaaaag aacaagctaa ggattagccg tcagcattcc
     2761 tacgataaca ttgtcgacaa acctagggag ctagacctta gcaggccctc ccggagcata
     2821 agcctcaagg acagggaacg gcttctggag ggaaattttt acggcagcct gtttagtgtc
     2881 ccctcaagca aactctcggg gaaaaaaagc tcccttttcc cccaaggtct ggaggacagc
     2941 aagaggagca agtctctctt gccagaccac acctccgata accctttcct ccactcccac
     3001 agggatgacc aacgcttggt tattgggaga tgcccctcgg acccttacaa acactcgttg
     3061 ccatcccagg cggtgaatga cagctatctt cggtcgtcct tgaggtcaac ggcatcgtac
     3121 tgttccaggg acagtcgggg ccacaatgat gtgtatattt cggagcatgt tatgccttat
     3181 gctgcaaata agaataatat gtactctacc cccagggttt taaattcctg cagcaataga
     3241 cgcgtgtaca agaaaatgcc tagtatcgaa tctgatgttt gtacagacta caaagaccat
     3301 gacggtgatt ataaagatca tgacatcgac tacaaggatg acgatgacaa gtaactcgag
     3361 ataatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt aactatgttg
     3421 ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct attgcttccc
     3481 gtatggcttt cattttctcc tccttgtata aatcctggtt agttcttgcc acggcggaac
     3541 tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt
     3601 ccgtggtgtt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc atctagcttt
     3661 atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa
     3721 gttaacaaca acaattgcat tcattttatg tttcaggttc agggggagat gtgggaggtt
     3781 ttttaaagat ctggagaccg gtgtggcacg tgcggaccga gcggccgcag gaacccctag
     3841 tgatggagtt ggccactccc tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa
     3901 aggtcgcccg acgcccgggc tttgcccggg cggcctcagt gagcgagcga gcgcgcagct
     3961 gcctgcaggg gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc
     4021 gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     4081 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ttagcgcccg ctcctttcgc
     4141 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     4201 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgattt
     4261 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     4321 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaactctat
     4381 ctcgggctat tcttttgatt tataagggat tttgccgatt tcggtctatt ggttaaaaaa
     4441 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt
     4501 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc
     4561 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     4621 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     4681 cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat
     4741 ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     4801 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     4861 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
     4921 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
     4981 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
     5041 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
     5101 tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg
     5161 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
     5221 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
     5281 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
     5341 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
     5401 aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt
     5461 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
     5521 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
     5581 atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa
     5641 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
     5701 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
     5761 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     5821 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     5881 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     5941 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     6001 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     6061 tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     6121 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     6181 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     6241 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     6301 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     6361 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     6421 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     6481 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     6541 cttttgctgg ccttttgctc acatgt
//
