LOCUS       sequence-491867-        6705 bp    DNA     circular SYN 04-NOV-2025
DEFINITION  3' AAVLINK plasmid for CUX2 expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6705)
  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  JOURNAL   Unpublished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 6705)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 4, 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            "Unpublished"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'-CUX2-FLAG
FEATURES             Location/Qualifiers
     source          1..6705
                     /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             3425..3490
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(3555..3575)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   3968..4108
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3968..4097
                     /label=AAV2 ITR
     rep_origin      4183..4638
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4270..4289)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4480..4501
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4655..4674)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4774..4796
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4834..4852)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4920..5024
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5025..5093,5094..5885)
                     /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(5243..5262)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6056..6644
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6545..6564
                     /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 ggggcggagg acgaaccccc
     1501 caggacgggc gagctcaagg ctgagggcgc gacggccgag gcgggcgcgc ggctgcccta
     1561 ctacccggcc tacgtgccgc gcaccctgaa gcccaccgtg ccgccgctga cccccgagca
     1621 gtacgagctg tacatgtacc gtgaggtaga cacgctggag ctcacccgcc aggtcaagga
     1681 gaagctggcc aagaacggca tctgccagag gatcttcggg gagaaggtgc tgggcctgtc
     1741 acagggcagc gtgagcgaca tgctgtcccg gccgaagcca tggagcaagc tgacgcagaa
     1801 ggggcgggag cccttcatcc gcatgcagct gtggctctct gaccagctcg gccaggcagt
     1861 gggccagcag cctggtgcct cccaggccag tcccacagaa ccaaggtcct caccatcccc
     1921 accccccagc cccacagagc ctgagaagag ctcccaggag ccgttgagcc tgtccctgga
     1981 gagcagcaag gagaaccagc agccagaggg ccgctccagc tcctcgttga gcgggaagat
     2041 gtactcaggc agccaggccc cagggggcat ccaggagatc gtggccatgt cccccgagct
     2101 ggacacgtac tccatcacca agagggtgaa ggaggtcctc acagacaaca atctagggca
     2161 gcggctgttt ggggaaagca tcctgggtct gacacagggc tccgtgtctg acctgctgtc
     2221 ccggcccaaa ccctggcaca agctgagcct gaaggggcgg gagccttttg tccgcatgca
     2281 gctgtggctc aatgaccccc ataacgtgga gaagctgagg gatatgaaga agctggagaa
     2341 gaaagcctac ctgaaacgtc gctatggcct catcagcacc ggctcagaca gtgagtcccc
     2401 ggccacccgc tcagagtgcc ccagcccctg cctgcagccc caggacctga gcctcctgca
     2461 gatcaagaag ccccgggtgg tgctggcacc cgaggagaag gaggcactgc ggaaggccta
     2521 tcagctggaa ccctacccct cgcagcagac catcgagctc ctctccttcc agctcaacct
     2581 caagaccaac accgtcatca actggttcca caactacagg tcccggatgc gccgggagat
     2641 gttggtggag gggacccagg atgagccaga ccttgatcca agcgggggtc ctggaatcct
     2701 accgccaggc cactcccacc cagaccccac cccgcagagc cctgactctg agactgagga
     2761 ccagaagcca accgtgaagg aactggagct tcaggagggc cctgaggaga acagcacacc
     2821 cctgaccacc caggacaagg cccaagtgag gatcaagcag gaacagatgg aggaggatgc
     2881 tgaggaagag gcaggcagcc agccccagga ctcaggggag ctggacaaag gccaaggtcc
     2941 ccccaaagag gagcatcccg accctccggg taatgatgga ctcccaaaag tggctcccgg
     3001 gcccctcctt ccaggtggat ccaccccaga ctgtccctca cttcatcccc aacaggagag
     3061 tgaggccggg gagcgacttc acccggaccc tttaagtttt aagtcagcct cagagtcctc
     3121 acgctgcagc ctggaggtgt cactgaactc gccctcggcc gcctcctcac caggcctcat
     3181 gatgtctgtg tcacctgtcc cctcctcctc agctcccatc tccccatccc cacctggcgc
     3241 cccccctgcc aaagtgccga gtgccagccc cactgctgac atggctggag ccttgcaccc
     3301 cagtgccaag gtgaacccca acttgcagcg gcggcatgag aagatggcca atctgaacaa
     3361 catcatttac cgactagagc gggctgccaa tcgggaggag gccctggagt gggagttctg
     3421 tacagactac aaagaccatg acggtgatta taaagatcat gacatcgact acaaggatga
     3481 cgatgacaag taactcgaga taatcaacct ctggattaca aaatttgtga aagattgact
     3541 ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg
     3601 tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa atcctggtta
     3661 gttcttgcca cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg
     3721 ctgttgggca ctgacaattc cgtggtgttt atttgtgaaa tttgtgatgc tattgcttta
     3781 tttgtaacca tctagcttta tttgtgaaat ttgtgatgct attgctttat ttgtaaccat
     3841 tataagctgc aataaacaag ttaacaacaa caattgcatt cattttatgt ttcaggttca
     3901 gggggagatg tgggaggttt tttaaagatc tggagaccgg tgtggcacgt gcggaccgag
     3961 cggccgcagg aacccctagt gatggagttg gccactccct ctctgcgcgc tcgctcgctc
     4021 actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg
     4081 agcgagcgag cgcgcagctg cctgcagggg cgcctgatgc ggtattttct ccttacgcat
     4141 ctgtgcggta tttcacaccg catacgtcaa agcaaccata gtacgcgccc tgtagcggcg
     4201 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgcct
     4261 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     4321 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
     4381 accccaaaaa acttgatttg ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
     4441 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
     4501 gaacaacact caactctatc tcgggctatt cttttgattt ataagggatt ttgccgattt
     4561 cggtctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa
     4621 tattaacgtt tacaatttta tggtgcactc tcagtacaat ctgctctgat gccgcatagt
     4681 taagccagcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
     4741 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
     4801 caccgtcatc accgaaacgc gcgagacgaa agggcctcgt gatacgccta tttttatagg
     4861 ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc
     4921 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     4981 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt
     5041 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
     5101 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg
     5161 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa
     5221 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc
     5281 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag
     5341 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa
     5401 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc
     5461 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg
     5521 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa
     5581 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa
     5641 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg
     5701 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag
     5761 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg
     5821 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt
     5881 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt
     5941 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac
     6001 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
     6061 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
     6121 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca
     6181 gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga
     6241 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca
     6301 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
     6361 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     6421 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     6481 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     6541 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     6601 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg
     6661 cctttttacg gttcctggcc ttttgctggc cttttgctca catgt
//
