LOCUS       sequence-491863-        6403 bp    DNA     circular SYN 04-NOV-2025
DEFINITION  3' AAVLINK plasmid for PXDN expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6403)
  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  JOURNAL   Unpublished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 6403)
  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            "Unpublished"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'-PXDN-FLAG
FEATURES             Location/Qualifiers
     source          1..6403
                     /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             3111..3176
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(3253..3273)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   3666..3806
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3666..3795
                     /label=AAV2 ITR
     rep_origin      3881..4336
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3968..3987)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4178..4199
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4353..4372)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4472..4494
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4532..4550)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4618..4722
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4723..4791,4792..5583)
                     /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(4941..4960)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5754..6342
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6243..6262
                     /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 gggcatcgtg cagcggtccg
     1501 ggaagccgct gctccccttc gccaccgggc cgcccacgga gtgcatgcgg gacgagaacg
     1561 agagccccat cccctgcttc ctggccgggg accaccgcgc caacgagcag ctgggcctga
     1621 ccagcatgca cacgctgtgg ttccgcgagc acaaccgcat tgccacggag ctgctcaagc
     1681 tgaacccgca ctgggacggc gacaccatct actatgagac caggaagatc gtgggtgcgg
     1741 agatccagca catcacctac cagcactggc tcccgaagat cctgggggag gtgggcatga
     1801 ggacgctggg agagtaccac ggctacgacc ccggcatcaa tgctggcatc ttcaacgcct
     1861 tcgccaccgc ggccttcagg tttggccaca cgcttgtcaa cccactgctt taccggctgg
     1921 acgagaactt ccagcccatt gcacaagatc acctccccct tcacaaagct ttcttctctc
     1981 ccttccggat tgtgaatgag ggcggcatcg atccgcttct cagggggctg ttcggggtgg
     2041 cggggaaaat gcgtgtgccc tcgcagctgc tgaacacgga gctcacggag cggctgttct
     2101 ccatggcaca cacggtggct ctggacctgg cggccatcaa catccagcgg ggccgggacc
     2161 acgggatccc accctaccac gactacaggg tctactgcaa tctatcggcg gcacacacgt
     2221 tcgaggacct gaaaaatgag attaaaaacc ctgagatccg ggagaaactg aaaaggttgt
     2281 atggctcgac actcaacatc gacctgtttc cggcgctcgt ggtggaggac ctggtgcctg
     2341 gcagccggct gggccccacc ctgatgtgtc ttctcagcac acagttcaag cgcctgcgag
     2401 atggggacag gttgtggtat gagaaccctg gggtgttctc cccggcccag ctgactcaga
     2461 tcaagcagac gtcgctggcc aggatcctat gcgacaacgc ggacaacatc acccgggtgc
     2521 agagcgacgt gttcagggtg gcggagttcc ctcacggcta cggcagctgt gacgagatcc
     2581 ccagggtgga cctccgggtg tggcaggact gctgtgaaga ctgtaggacc agggggcagt
     2641 tcaatgcctt ttcctatcat ttccgaggca gacggtctct tgagttcagc taccaggagg
     2701 acaagccgac caagaaaaca agaccacgga aaatacccag tgttgggaga cagggggaac
     2761 atctcagcaa cagcacctca gccttcagca cacgctcaga tgcatctggg acaaatgact
     2821 tcagagagtt tgttctggaa atgcagaaga ccatcacaga cctcagaaca cagataaaga
     2881 aacttgaatc acggctcagt accacagagt gcgtggatgc cgggggcgaa tctcacgcca
     2941 acaacaccaa gtggaaaaaa gatgcatgca ccatttgtga atgcaaagac gggcaggtca
     3001 cctgcttcgt ggaagcttgc ccccctgcca cctgtgctgt ccccgtgaac atcccagggg
     3061 cctgctgtcc agtctgctta cagaagaggg cggaggaaaa gccctgtaca gactacaaag
     3121 accatgacgg tgattataaa gatcatgaca tcgactacaa ggatgacgat gacaagtaag
     3181 aattctgatg actcgagata atcaacctct ggattacaaa atttgtgaaa gattgactgg
     3241 tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta
     3301 tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttagt
     3361 tcttgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct
     3421 gttgggcact gacaattccg tggtgtttat ttgtgaaatt tgtgatgcta ttgctttatt
     3481 tgtaaccatc tagctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta
     3541 taagctgcaa taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg
     3601 gggagatgtg ggaggttttt taaagatctg gagaccggtg tggcacgtgc ggaccgagcg
     3661 gccgcaggaa cccctagtga tggagttggc cactccctct ctgcgcgctc gctcgctcac
     3721 tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag
     3781 cgagcgagcg cgcagctgcc tgcaggggcg cctgatgcgg tattttctcc ttacgcatct
     3841 gtgcggtatt tcacaccgca tacgtcaaag caaccatagt acgcgccctg tagcggcgca
     3901 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcctta
     3961 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     4021 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     4081 cccaaaaaac ttgatttggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     4141 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     4201 acaacactca actctatctc gggctattct tttgatttat aagggatttt gccgatttcg
     4261 gtctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata
     4321 ttaacgttta caattttatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     4381 agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg
     4441 gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca
     4501 ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt
     4561 aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc
     4621 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     4681 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     4741 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     4801 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     4861 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     4921 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     4981 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     5041 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     5101 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     5161 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     5221 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     5281 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     5341 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     5401 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     5461 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     5521 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     5581 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     5641 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     5701 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     5761 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     5821 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     5881 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     5941 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     6001 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     6061 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     6121 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     6181 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     6241 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     6301 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     6361 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgt
//
