LOCUS       sequence-476860-        6895 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  3' AAVLINK plasmid for KDM6A expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6895)
  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  JOURNAL   Unpublished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 6895)
  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            "Unpublished"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' KDM6A
FEATURES             Location/Qualifiers
     source          1..6895
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(132..1163)
                     /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   1310..1439
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(1501..2089)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1581..1600)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2260..3051,3052..3120))
                     /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     2883..2902
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3121..3225)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3293..3311
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3349..3371)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3471..3490
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(3507..3962)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3644..3665)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3856..3875
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   4037..4177
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4048..4177
                     /label=AAV2 ITR
     primer_bind     4570..4590
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4655..4720)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
ORIGIN
        1 gtgcctatca gaaacgcaag agtcttctct gtctcgacaa gcccagtttc ataaattgct
       61 ataaagtatc ctatacgaag ttatactagt tattggtctc cttaaacctg tcttgtaacc
      121 ttgatactta cctaatcgcc atcttccagc aggcgcacca ttgcccctgt ttcactatcc
      181 aggttacgga tatagttcat tacaatattt acattggtcc agccaccagc ttgcatgatc
      241 tccggtattg aaactccagc gcgggccata tctcgcgcgg ctccgacacg ggcactgtgt
      301 ccagaccagg ccaggtatct ctgaccagag tcatccttag cgccgtaaat caatcgatga
      361 gttgcttcaa aaatcccttc cagggcgcga gttgatagct ggctggtggc agatggcgcg
      421 gcaacaccat tttttctgac ccggcaaaac aggtagttat tcggatcatc agctacacca
      481 gagacggaaa tccatcgctc gaccagttta gttaccccca ggctaagtgc cttctctaca
      541 cctgcggtgc taaccagcgt tttcgttctg ccaatatgga ttaacattct cccaccgtca
      601 gtacgtgaga tatctttaac cctgatcctg gcaatttcgg ctatacgtaa cagggtgtta
      661 taagcaatcc ccagaaatgc cagattacgt atatcctggc agcgatcgct attttccatg
      721 agtgaacgaa cctggtcgaa atcagtgcgt tcgaacgcta gagcctgttt tgcacgttca
      781 ccggcatcaa cgttttcttt tcggatccgc cgcataacca gtgaaacagc attgctgtca
      841 cttggtcgtg gcagcccgga ccgacgatga agcatgttta gctggcccaa atgttgctgg
      901 atggttttta ctgccagacc gcgcgcctga agatatagaa gataatcgcg aacatcttca
      961 ggttctgcgg gaaaccattt ccggttattc aacttgcacc atgccgccca cgaccggcaa
     1021 acggacagaa gcattttcca ggtatgctca gaaaacgcct ggcgatccct gaacatgtcc
     1081 atcaggttct tgcgaacctc atcactcgtt gcatcgaccg gtaatgcagg caaattttgg
     1141 tgtacggtca gtaaattgga catggtggcg tacgagtaac ggcacggcgt ccacgacact
     1201 cccagcacca ctgccggagc gcgagtctgc gccccgcgat atatatatgg tgcgccgtat
     1261 ccctccgccg cggcggagcc gcctcgcgca gagcgacgcg tgcggccgca ggaaccccta
     1321 gtgatggagt tggccactcc ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca
     1381 aaggtcgccc gacgcccggg cggcctcagt gagcgagcga gcgcgcagct gcctgcagga
     1441 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     1501 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     1561 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     1621 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     1681 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     1741 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     1801 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     1861 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     1921 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     1981 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     2041 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     2101 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     2161 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     2221 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     2281 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     2341 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     2401 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     2461 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     2521 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     2581 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     2641 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     2701 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     2761 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     2821 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     2881 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     2941 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     3001 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     3061 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     3121 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     3181 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     3241 agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg
     3301 tatcacgagg ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat
     3361 gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg
     3421 tcagggcgcg tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga
     3481 gcagattgta ctgagagtgc accataaaat tgtaaacgtt aatattttgt taaaattcgc
     3541 gttaaatttt tgttaaatca gctcattttt taaccaatag accgaaatcg gcaaaatccc
     3601 ttataaatca aaagaatagc ccgagataga gttgagtgtt gttccagttt ggaacaagag
     3661 tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga
     3721 tggcccacta cgtgaaccat cacccaaatc aagttttttg gggtcgaggt gccgtaaagc
     3781 actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa
     3841 cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctaaggcgc tggcaagtgt
     3901 agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc
     3961 gtactatggt tgctttgacg tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa
     4021 taccgcatca ggcgcccctg caggcagctg cgcgctcgct cgctcactga ggccgcccgg
     4081 gcaaagcccg ggcgtcgggc gacctttggt cgcccggcct cagtgagcga gcgagcgcgc
     4141 agagagggag tggccaactc catcactagg ggttcctgcg gccgctcggt ccgcacgtgc
     4201 cacaccggtc tccagatctt taaaaaacct cccacatctc cccctgaacc tgaaacataa
     4261 aatgaatgca attgttgttg ttaacttgtt tattgcagct tataatggtt acaaataaag
     4321 caatagcatc acaaatttca caaataaagc tagatggtta caaataaagc aatagcatca
     4381 caaatttcac aaataaacac cacggaattg tcagtgccca acagccgagc ccctgtccag
     4441 cagcgggcaa ggcaggcggc gatgagttcc gccgtggcaa gaactaacca ggatttatac
     4501 aaggaggaga aaatgaaagc catacgggaa gcaatagcat gatacaaagg cattaaagca
     4561 gcgtatccac atagcgtaaa aggagcaaca tagttaagaa taccagtcaa tctttcacaa
     4621 attttgtaat ccagaggttg attatctcga gttacttgtc atcgtcatcc ttgtagtcga
     4681 tgtcatgatc tttataatca ccgtcatggt ctttgtagtc tgtacaagat gaggcggatg
     4741 gtaatggagg agctaatgta aattggtcat agacttgcat caggtcctcc attttgtact
     4801 gttctagcac cacaaagttt tccaagtttc cgcttgtttt tcgtgcacaa tcttggcaat
     4861 gtactatgta ggtctttcgt gaattactct cattagtgac aaaaagcaga tcaaaaacct
     4921 ccacttcaca aatgctacag taatgagctg gttcttcttt tgtccgccca tgccatataa
     4981 tctcttttcc tgcagcaatg agagcttccc tcaatgtctg acattgcttc agagttctta
     5041 gaagacaata cttaatcatt tcaaaaagct ttggatctga gaccttgata tttcgtgcca
     5101 tattccagga aagatgaacc atgggtacta ttgacttcac actttgcaat ttgttccatt
     5161 cgtaccgttc cactgccaat ttatactggc aggctgtaag tggaccaaca ttccaagcaa
     5221 tgttgttgca ccagccaata gcctgaaccc aatgaacagt gcctgcattt atccagacca
     5281 aatctccagg tcgctgaata aacctataca ctggaacatt tgcttcataa agatcttcaa
     5341 gattgggcca ccaagaaccc attaggaaat tcaaattatt tttttcacag aagtcattca
     5401 gaacacccca gtaaccttca ggaacaacaa accattcaca gtcacctggg ccaatattta
     5461 tgttaactga acagaagttg ttattttcct gatgacctgg tgttctgctc cctggaactt
     5521 tcatgtatag ttgaactgtg ttcatgccca atatggtatg accaacatgg cttagaagat
     5581 ttcctgctga tacgacacgc acaaaagcag gaagtttagt cagctcatgt agctgcaact
     5641 tccacttttt gtcatcagat aggtcaatat tggtcccaaa ctttatggtt ttaaagggtc
     5701 ctttcctcct cctcccagaa ttatctgacg atgtagattc actatctgag tggtctttat
     5761 gatgacttct tttttcattt tcttctctca atgattcctg gaatgaggag gcctggtact
     5821 gtgcatattt agcaattgta gtatgagatc tattactttc acaatgccag attttctttg
     5881 ttccagtggg atcccagttt tcatctgctg gctgcaacaa ctgtgtcctc acttctacca
     5941 tatgttcatt gttagcttcc accaaagttt tagtagagaa aagtcccagg tctaacttaa
     6001 gagctccagc aaggccacgt attactgtaa cagggttgtt cggatttgta caaaattgat
     6061 gtaatggagg aaagaaagca tcacgtttat tttccaagta aatactaggt gtaggtggat
     6121 tcaacttgtc ctttggcaag ggagggtatg gtgaagatgg tggtcttgga ggtggacatt
     6181 tatccaacaa aatgctactg ttagataagc catttttacc tagattcctg catgccttca
     6241 gaacttctgc tgagctgggg tatatggaca cagacattga tggtatgatc tgtggactag
     6301 gtttgtggtt aaccagaagc agatctgttt tcatggggct ctgagattct tccatccctt
     6361 ctccattaat tgtgtgtagc ccactgtgag ggctgttaag gctggtaaca ctgtttgtgg
     6421 ttgtctgctc agtggatttt ggagaaggtg ttgctgttga aatggctgaa gatggtgaag
     6481 aggcaacaga gttatcagtc tttgtatgaa cagctgggtg gatgttattg actttgtcac
     6541 aggttccagt acccacattg ttattggctt ttcccatcaa caaggcagag agctgaggat
     6601 tgtctgaact tagtaaacct ggtgacttag aatctccatg gctaggactg caaacagcat
     6661 ctgccgtcat ctgatggaca tgattaggaa gtccctcgac actggcagtg ctgttaggtg
     6721 tctctccagt gtgcctgctt gtttcaggca ccgtcaatat gtttcctgaa ggcttgctct
     6781 ctttggttaa ggtaatgcct tgttgtccac ctgaggtagc agtgtgagag gagaggtgat
     6841 tgagagcagc cccctgtgga gagaaaggca aagtggatgt cagtaagacc aatag
//
