LOCUS       sequence-476875-        6723 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  3' AAVLINK plasmid for KDIP2A expression .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6723)
  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  JOURNAL   Unpublished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 6723)
  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            "Unpublished"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' DIP2A
FEATURES             Location/Qualifiers
     source          1..6723
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1964..2029
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(2094..2114)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   2507..2647
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2507..2636
                     /label=AAV2 ITR
     rep_origin      2722..3177
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2809..2828)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3019..3040
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3194..3213)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3313..3335
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3373..3391)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3459..3563
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3564..3632,3633..4424)
                     /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(3782..3801)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4595..5183
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5084..5103
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   5245..5374
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     CDS             5521..6552
                     /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"
ORIGIN
        1 agggacgctg cacccgtgta atgtgctgat gtgccctcac acctgtgtta ccaacctccc
       61 caaacctcgt cagaaacaac cagaggttgg accagcctca atgatcgtgg ggaacctggt
      121 tgctgggaag agaatcgctc aggcttccgg gagagagctc gcccacctgg aggacagcga
      181 ccaggcacgg aagttcctgt tcctggctga cgtgctgcag tggcgtgccc acaccactcc
      241 tgaccacccg ctgttcttgc tgctgaacgc caagggcacc gtcacaagca ctgcaacctg
      301 tgtccagctg cacaaaaggg ctgagagagt ggccgcggct ctgatggaga agggaagact
      361 gagtgttggg gaccatgtgg ctctggtcta cccaccaggg gtggacctca ttgccgcgtt
      421 ctatggctgc ttgtactgtg gctgcgtgcc tgtcaccgtg cggcccccgc accctcagaa
      481 cctcggcacc acactgccca ccgtcaagat gatcgtggag gtcagcaagt ctgcatgcgt
      541 cctcaccacg caggctgtca cacggctgct caggtccaag gaggctgctg ctgccgtgga
      601 catcaggacc tggcccacca tcctagacac agatgacatc ccaaaaaaga agatagcaag
      661 cgttttcagg cccccctccc ccgatgtcct cgcatacttg gacttcagcg tgtcaaccac
      721 tgggatatta gcgggagtga agatgtcgca cgcggccaca agcgccttat gccgctccat
      781 aaagctgcag tgtgagctgt acccctcgcg gcagatcgcc atctgcctcg acccctactg
      841 tggccttggt tttgccctgt ggtgtctgtg cagtgtctac tcgggacacc aatcagtgct
      901 ggtgcccccg ctggagctgg agagcaacgt gtccctgtgg ctgtcggccg tcagccagta
      961 caaggcccgc gtcaccttct gctcctactc tgtgatggag atgtgcacca agggcctagg
     1021 cgcacagacg ggtgtcctca ggatgaaggg ggtgaacctg tcatgtgtgc gcacgtgcat
     1081 ggtggtcgcc gaggagcggc ccaggattgc gctgacccag tccttctcca agctcttcaa
     1141 ggacctgggc ctgccggccc gcgccgtaag caccacgttc gggtgcaggg tcaacgtggc
     1201 catctgcctc cagggcacag ctggcccgga ccccacaacc gtctacgtgg acatgcgggc
     1261 actgcgccat gacagggttc gtttggtaga acggggttct ccgcacagcc tgccattgat
     1321 ggagtctgga aagatcctcc ccggcgtgaa ggtcatcatc gcacacaccg agaccaaagg
     1381 acccttggga gactcacacc tgggagagat ctgggtaagc agcccccaca atgccaccgg
     1441 gtactacacc gtttacgggg aggaggcgct tcatgccgac cacttcagtg cccggctgag
     1501 ttttggagac acacagacca tctgggcaag gaccggctac cttggcttcc ttcggcgaac
     1561 agagctcact gatgccagtg gagggcggca cgatgcactg tatgtggttg ggtctctgga
     1621 tgaaactctg gagctcagag gcatgcggta ccaccccatc gacattgaga cctctgtcat
     1681 ccgagcacac aggagcatcg ctgagtgtgc cgtattcacc tggaccaacc tgctggtggt
     1741 ggtggtggag ctggatgggc tagagcagga tgccctggac ctggtggccc tggtgaccaa
     1801 cgtggtgctg gaggagcact acctggtcgt gggagtggtg gtcatcgtgg acccaggggt
     1861 gatccctatc aactctcggg gtgagaagca gcgcatgcac ctgcgggacg gcttcctggc
     1921 tgaccagctg gaccccatct atgtcgccta caacatgtgt acagactaca aagaccatga
     1981 cggtgattat aaagatcatg acatcgacta caaggatgac gatgacaagt aactcgagat
     2041 aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct
     2101 ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt
     2161 atggctttca ttttctcctc cttgtataaa tcctggttag ttcttgccac ggcggaactc
     2221 atcgccgcct gccttgcccg ctgctggaca ggggctcggc tgttgggcac tgacaattcc
     2281 gtggtgttta tttgtgaaat ttgtgatgct attgctttat ttgtaaccat ctagctttat
     2341 ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt
     2401 taacaacaac aattgcattc attttatgtt tcaggttcag ggggagatgt gggaggtttt
     2461 ttaaagatct ggagaccggt gtggcacgtg cggaccgagc ggccgcagga acccctagtg
     2521 atggagttgg ccactccctc tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag
     2581 gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc gcgcagctgc
     2641 ctgcaggggc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
     2701 atacgtcaaa gcaaccatag tacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg
     2761 tggttacgcg cagcgtgacc gctacacttg ccagcgcctt agcgcccgct cctttcgctt
     2821 tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc
     2881 tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgatttgg
     2941 gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg
     3001 agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc aactctatct
     3061 cgggctattc ttttgattta taagggattt tgccgatttc ggtctattgg ttaaaaaatg
     3121 agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgttt acaattttat
     3181 ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagccc cgacacccgc
     3241 caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag
     3301 ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg
     3361 cgagacgaaa gggcctcgtg atacgcctat ttttataggt taatgtcatg ataataatgg
     3421 tttcttagac gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     3481 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     3541 aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct
     3601 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag
     3661 atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta
     3721 agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc
     3781 tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca
     3841 tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg
     3901 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg
     3961 ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca
     4021 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa
     4081 acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa
     4141 ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg gaggcggata
     4201 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat
     4261 ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc
     4321 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata
     4381 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt
     4441 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga
     4501 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
     4561 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     4621 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     4681 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     4741 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     4801 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     4861 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     4921 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     4981 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     5041 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     5101 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     5161 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     5221 tttgctggcc ttttgctcac atgtcctgca ggcagctgcg cgctcgctcg ctcactgagg
     5281 ccgcccgggc gtcgggcgac ctttggtcgc ccggcctcag tgagcgagcg agcgcgcaga
     5341 gagggagtgg ccaactccat cactaggggt tcctgcggcc gcacgcgtcg ctctgcgcga
     5401 ggcggctccg ccgcggcgga gggatacggc gcaccatata tatatcgcgg ggcgcagact
     5461 cgcgctccgg cagtggtgct gggagtgtcg tggacgccgt gccgttactc gtacgccacc
     5521 atgtccaatt tactgaccgt acaccaaaat ttgcctgcat taccggtcga tgcaacgagt
     5581 gatgaggttc gcaagaacct gatggacatg ttcagggatc gccaggcgtt ttctgagcat
     5641 acctggaaaa tgcttctgtc cgtttgccgg tcgtgggcgg catggtgcaa gttgaataac
     5701 cggaaatggt ttcccgcaga acctgaagat gttcgcgatt atcttctata tcttcaggcg
     5761 cgcggtctgg cagtaaaaac catccagcaa catttgggcc agctaaacat gcttcatcgt
     5821 cggtccgggc tgccacgacc aagtgacagc aatgctgttt cactggttat gcggcggatc
     5881 cgaaaagaaa acgttgatgc cggtgaacgt gcaaaacagg ctctagcgtt cgaacgcact
     5941 gatttcgacc aggttcgttc actcatggaa aatagcgatc gctgccagga tatacgtaat
     6001 ctggcatttc tggggattgc ttataacacc ctgttacgta tagccgaaat tgccaggatc
     6061 agggttaaag atatctcacg tactgacggt gggagaatgt taatccatat tggcagaacg
     6121 aaaacgctgg ttagcaccgc aggtgtagag aaggcactta gcctgggggt aactaaactg
     6181 gtcgagcgat ggatttccgt ctctggtgta gctgatgatc cgaataacta cctgttttgc
     6241 cgggtcagaa aaaatggtgt tgccgcgcca tctgccacca gccagctatc aactcgcgcc
     6301 ctggaaggga tttttgaagc aactcatcga ttgatttacg gcgctaagga tgactctggt
     6361 cagagatacc tggcctggtc tggacacagt gcccgtgtcg gagccgcgcg agatatggcc
     6421 cgcgctggag tttcaatacc ggagatcatg caagctggtg gctggaccaa tgtaaatatt
     6481 gtaatgaact atatccgtaa cctggatagt gaaacagggg caatggtgcg cctgctggaa
     6541 gatggcgatt aggtaagtat caaggttaca agacaggttt aaggagacca ataactagta
     6601 taacttcgta taggatactt tatagcaatt tatgaaactg ggcttgtcga gacagagaag
     6661 actcttgcgt ttctgatagg cacctattgg tcttactgac atccactttg cctttctctc
     6721 cac
//
