LOCUS       sequence-476856-        6319 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  3' AAVLINK plasmid for PREX1 expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6319)
  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  JOURNAL   Unpublished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 6319)
  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            "Unpublished"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' PREX1
FEATURES             Location/Qualifiers
     source          1..6319
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(603..1394,1395..1463))
                     /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     1226..1245
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1464..1568)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1636..1654
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(1692..1714)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     1814..1833
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(1850..2305)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1987..2008)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2199..2218
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   2380..2520
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2391..2520
                     /label=AAV2 ITR
     primer_bind     2913..2933
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3010..3075)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(4619..4638)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             complement(4794..5825)
                     /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"
     repeat_region   5972..6101
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(join(6163..6319,1..432))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6243..6262)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
       61 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
      121 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
      181 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
      241 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
      301 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
      361 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
      421 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
      481 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
      541 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
      601 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
      661 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
      721 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
      781 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
      841 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
      901 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
      961 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     1021 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     1081 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     1141 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     1201 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     1261 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     1321 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     1381 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     1441 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     1501 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     1561 gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg
     1621 acattaacct ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat
     1681 gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg
     1741 gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc
     1801 tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccataa aattgtaaac
     1861 gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
     1921 tagaccgaaa tcggcaaaat cccttataaa tcaaaagaat agcccgagat agagttgagt
     1981 gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg
     2041 cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccaa atcaagtttt
     2101 ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga
     2161 gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg
     2221 ggcgctaagg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg
     2281 cttaatgcgc cgctacaggg cgcgtactat ggttgctttg acgtatgcgg tgtgaaatac
     2341 cgcacagatg cgtaaggaga aaataccgca tcaggcgccc ctgcaggcag ctgcgcgctc
     2401 gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt ggtcgcccgg
     2461 cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact aggggttcct
     2521 gcggccgctc ggtccgcacg tgccacaccg gtctccagat ctttaaaaaa cctcccacat
     2581 ctccccctga acctgaaaca taaaatgaat gcaattgttg ttgttaactt gtttattgca
     2641 gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa agctagatgg
     2701 ttacaaataa agcaatagca tcacaaattt cacaaataaa caccacggaa ttgtcagtgc
     2761 ccaacagccg agcccctgtc cagcagcggg caaggcaggc ggcgatgagt tccgccgtgg
     2821 caagaactaa ccaggattta tacaaggagg agaaaatgaa agccatacgg gaagcaatag
     2881 catgatacaa aggcattaaa gcagcgtatc cacatagcgt aaaaggagca acatagttaa
     2941 gaataccagt caatctttca caaattttgt aatccagagg ttgattatct cgagtcatca
     3001 gaattcttac ttgtcatcgt catccttgta gtcgatgtca tgatctttat aatcaccgtc
     3061 atggtctttg tagtctgtac agaggtcccc atccaccggc ggctggcaga ggcggtagag
     3121 gcgcggagca ccctggggca tctggtcctt gactcgcagg tttttggcca gaatctccac
     3181 ccttgggccc tgcttccgca tgatgtccgt ggcctgcatg atgcacttgg gcagcaaccc
     3241 gtggctccgt gccaagatgg ccgcctgctc cagggacacg ctcagggtgc tcctctgcat
     3301 gcctgtgcca cacatgacca tctggcaggc ccccaggcgg tagcagagct ccgaggagat
     3361 ggggatgagg ccggcgccca cactcccagc agcaccaggc tttgggtgga caaaggactt
     3421 catgaggcgg cagagctcat ccagggcatt gatggggcgg atcagctggt ctatctttac
     3481 cgccgtggtg ctggcatccg tgggcaggtt agaccgctcc aggtaaaatg ccctgagttt
     3541 gcggtaatac tgctgaactt tctccaggga ctgcgcgttg atgtcctgct gcaaatcctc
     3601 cgcggcctgg ctgcctggag aaggcagccc ctccacgttc tccagcactt tcgtgaacag
     3661 cgctgtgtgc agcctcaggc tggccccacc ctccaggcgg gagggcagct tggagaagtg
     3721 gtagctgtcg aggtagaaga tgaccttcag cgcctgccgg ctgccctcaa tgtggtagaa
     3781 gacgttggtg ttggccacat agttctcgtc cagctgctta aaggagaagg tgacattgtc
     3841 cagctctgac agcgtcaccc agatgtcctc cagcatggtc cgttcctcct tcactgtggc
     3901 tggcgagagc agggactggc agtgcagcag gacgcccgtg gccgccacct gctccagcca
     3961 cttgcggctg gcgtcgcggc tgctctcctc gtactcgcca ttgttgttgt agcggtagcc
     4021 cagggccgcc agcagctgct cggagaaggt gcacacggcg gccaccaggg cctggcagaa
     4081 gatcgcgtct ctgcgcagct gcagctccgt gtctgtgcac ttcagcaagg ccaggagcag
     4141 ctggttcttc ccatcttcca catatctctg aatgttgtcc accagggtct tgatggagtt
     4201 ggggaggttc caggggtcct cctggatgct aatctggatc aggctccggc cacggattgt
     4261 acactcttct ttctgtttaa actcttgtaa gctgtggttc atagggaaat gcttggtctc
     4321 ttcgaaagcc cggctcatga ctggcccctt gaggagagca ttgatggagt ccacctggtt
     4381 aaagaggtgc tccaggcagc catgaagctt gtcctgcttg tcagatggga tccgcatgtc
     4441 acagggcagc tcatctccag accccatctc gtcgctgccc aagtaggagc tgttacttct
     4501 cacgctggtg taggacagga ccgagtctcg attgctgtta cactcgctgt aggagtcgcg
     4561 ataactcatg gtgtcgtggc ctgagtcctc ctggtcctcc tcggccacct tgaagcacac
     4621 ctgtggagag aaaggcaaag tggatgtcag taagaccaat aggtgcctat cagaaacgca
     4681 agagtcttct ctgtctcgac aagcccagtt tcataaattg ctataaagta tcctatacga
     4741 agttatacta gttattggtc tccttaaacc tgtcttgtaa ccttgatact tacctaatcg
     4801 ccatcttcca gcaggcgcac cattgcccct gtttcactat ccaggttacg gatatagttc
     4861 attacaatat ttacattggt ccagccacca gcttgcatga tctccggtat tgaaactcca
     4921 gcgcgggcca tatctcgcgc ggctccgaca cgggcactgt gtccagacca ggccaggtat
     4981 ctctgaccag agtcatcctt agcgccgtaa atcaatcgat gagttgcttc aaaaatccct
     5041 tccagggcgc gagttgatag ctggctggtg gcagatggcg cggcaacacc attttttctg
     5101 acccggcaaa acaggtagtt attcggatca tcagctacac cagagacgga aatccatcgc
     5161 tcgaccagtt tagttacccc caggctaagt gccttctcta cacctgcggt gctaaccagc
     5221 gttttcgttc tgccaatatg gattaacatt ctcccaccgt cagtacgtga gatatcttta
     5281 accctgatcc tggcaatttc ggctatacgt aacagggtgt tataagcaat ccccagaaat
     5341 gccagattac gtatatcctg gcagcgatcg ctattttcca tgagtgaacg aacctggtcg
     5401 aaatcagtgc gttcgaacgc tagagcctgt tttgcacgtt caccggcatc aacgttttct
     5461 tttcggatcc gccgcataac cagtgaaaca gcattgctgt cacttggtcg tggcagcccg
     5521 gaccgacgat gaagcatgtt tagctggccc aaatgttgct ggatggtttt tactgccaga
     5581 ccgcgcgcct gaagatatag aagataatcg cgaacatctt caggttctgc gggaaaccat
     5641 ttccggttat tcaacttgca ccatgccgcc cacgaccggc aaacggacag aagcattttc
     5701 caggtatgct cagaaaacgc ctggcgatcc ctgaacatgt ccatcaggtt cttgcgaacc
     5761 tcatcactcg ttgcatcgac cggtaatgca ggcaaatttt ggtgtacggt cagtaaattg
     5821 gacatggtgg cgtacgagta acggcacggc gtccacgaca ctcccagcac cactgccgga
     5881 gcgcgagtct gcgccccgcg atatatatat ggtgcgccgt atccctccgc cgcggcggag
     5941 ccgcctcgcg cagagcgacg cgtgcggccg caggaacccc tagtgatgga gttggccact
     6001 ccctctctgc gcgctcgctc gctcactgag gccgggcgac caaaggtcgc ccgacgcccg
     6061 ggcggcctca gtgagcgagc gagcgcgcag ctgcctgcag gacatgtgag caaaaggcca
     6121 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     6181 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     6241 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     6301 gccgcttacc ggatacctg
//
