LOCUS       sequence-491423-        6988 bp    DNA     circular SYN 03-NOV-2025
DEFINITION  3' AAVLINK plasmid for NRXN2 expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6988)
  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  JOURNAL   Unpublished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 6988)
  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            "Unpublished"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'-NRXN2-FLAG
FEATURES             Location/Qualifiers
     source          1..6988
                     /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             3708..3773
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(3838..3858)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   4251..4391
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4251..4380
                     /label=AAV2 ITR
     rep_origin      4466..4921
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4553..4572)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4763..4784
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4938..4957)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5057..5079
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5117..5135)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5203..5307
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5308..5376,5377..6168)
                     /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(5526..5545)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6339..6927
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6828..6847
                     /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 ggggtacatc cactacgtgt
     1501 ttgacctggg gaatggcccg tccttgatga aggggaactc agacaaacca gtcaatgaca
     1561 accagtggca caacgtggtg gtgtccaggg acccaggcaa cgtgcacacg ctcaagattg
     1621 actcccgcac tgtcacgcag cactccaatg gcgcccgaaa cctcgatctc aaaggggagt
     1681 tgtacattgg cggtctgagc aagaatatgt tcagcaacct gcccaagctg gtggcctccc
     1741 gggatggctt tcagggctgc ctggcctcag tggacctcaa cggacgtctc ccagacctca
     1801 tcgccgacgc cctgcaccgc attgggcagg tggagagggg ctgtgatggc cccagcacca
     1861 cctgcactga agagtcctgt gccaaccagg gcgtctgctt gcagcagtgg gatggcttca
     1921 cctgcgactg caccatgact tcctatggag gccctgtctg caatgatccc gggaccacat
     1981 acatctttgg gaagggggga gcgctcatca cctacacgtg gccccccaat gacaggccca
     2041 gcacgaggat ggatcgcctg gccgtgggct tcagcaccca ccagcggagc gctgtgctgg
     2101 tgcgggtgga cagcgcctcc ggccttggag actacctgca gctgcacatc gaccagggca
     2161 ccgtgggggt gatctttaac gtgggcacgg acgacattac catcgacgag cccaacgcca
     2221 tagtaagcga cggcaaatac cacgtggtgc gcttcactcg aagcggcggc aacgccaccc
     2281 tgcaggtgga cagctggccg gtcaacgagc ggtacccggc aggaaacttt gataacgagc
     2341 gcctggcgat tgctagacag agaatcccct accggcttgg tcgagtagta gatgagtggc
     2401 tgctcgacaa aggccgccag ctgaccatct tcaacagcca ggctgccatc aagatcgggg
     2461 gccgggatca gggccgcccc ttccagggcc aggtgtccgg cctctactac aatgggctca
     2521 aggtgctggc gctggccgcc gagagcgacc ccaatgtgcg gactgagggt cacctgcgcc
     2581 tggtggggga ggggccgtcc gtgctgctca gtgcggagac cacggccacc accctgctgg
     2641 ctgacatggc caccaccatc atggagacta ccaccaccat ggccactacc accacgcgcc
     2701 ggggccgctc ccccacactg agggacagca ccacccagaa cacagatgac ctgctggtgg
     2761 cctctgctga gtgtccaagc gatgatgagg acctggagga gtgtgagccc agtactggag
     2821 gagagttaat attgcccatt atcacggagg actccttaga cccccctccc gtggccaccc
     2881 gatccccctt cgtgcccccg ccccctacct tctacccctt cctcacggga gtgggcgcca
     2941 cccaagacac gctgcccccg cccgccgcgc gccgcccgcc ctctgggggc ccgtgccagg
     3001 ccgagcggga cgacagcgac tgcgaggagc ccatcgaggc ctcgggcttc gcctccgggg
     3061 aggtctttga ctccagcctc ccccccacgg acgacgagga cttttacacc acctttcccc
     3121 tggtcacgga ccgcaccacc ctcctgtcac cccgcaaacc cgctccccgg cccaacctca
     3181 ggacagatgg ggccacgggc gcccctgggg tgctgtttgc cccctccgcc ccggccccca
     3241 acctgccggc gggcaaaatg aaccaccgag acccgcttca gcccttgctg gagaacccgc
     3301 ccttggggcc cggggccccc acgtcctttg agccgcggag gccccctccc ctgcgccccg
     3361 gcgtgacctc agcccccggc ttcccccatc tgcccacagc caaccccaca gggcctgggg
     3421 agcggggccc gccgggcgca gtggaggtga tccgggagtc cagcagcacc acgggcatgg
     3481 tggtgggcat tgtggcggcg gcggcgctct gcatcctcat cctcctctac gccatgtata
     3541 agtaccgcaa tcgtgatgag ggctcctacc aggtggacca gagccgaaac tacatcagta
     3601 actcggccca gagcaatggg gcggtggtga aagagaaggc cccggctgcc cccaagacgc
     3661 ccagcaaggc caagaagaac aaagacaagg agtattatgt ctgtacagac tacaaagacc
     3721 atgacggtga ttataaagat catgacatcg actacaagga tgacgatgac aagtaactcg
     3781 agataatcaa cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt
     3841 tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc
     3901 ccgtatggct ttcattttct cctccttgta taaatcctgg ttagttcttg ccacggcgga
     3961 actcatcgcc gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa
     4021 ttccgtggtg tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccatctagct
     4081 ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac
     4141 aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag atgtgggagg
     4201 ttttttaaag atctggagac cggtgtggca cgtgcggacc gagcggccgc aggaacccct
     4261 agtgatggag ttggccactc cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc
     4321 aaaggtcgcc cgacgcccgg gctttgcccg ggcggcctca gtgagcgagc gagcgcgcag
     4381 ctgcctgcag gggcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca
     4441 ccgcatacgt caaagcaacc atagtacgcg ccctgtagcg gcgcattaag cgcggcgggt
     4501 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccttagcgcc cgctcctttc
     4561 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     4621 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     4681 ttgggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     4741 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaactct
     4801 atctcgggct attcttttga tttataaggg attttgccga tttcggtcta ttggttaaaa
     4861 aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac gtttacaatt
     4921 ttatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gccccgacac
     4981 ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga
     5041 caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa
     5101 cgcgcgagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata
     5161 atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     5221 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     5281 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     5341 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     5401 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     5461 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     5521 gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc
     5581 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     5641 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     5701 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     5761 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     5821 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta
     5881 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     5941 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     6001 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
     6061 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     6121 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     6181 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     6241 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     6301 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     6361 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     6421 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     6481 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     6541 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     6601 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     6661 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     6721 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     6781 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     6841 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     6901 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     6961 gccttttgct ggccttttgc tcacatgt
//
