LOCUS       sequence-491424-        7110 bp    DNA     circular SYN 03-NOV-2025
DEFINITION  3' AAVLINK plasmid for KDM3B expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7110)
  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  JOURNAL   Unpublished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 7110)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 3, 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            "Unpublished"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'-KDM3B-FLAG
FEATURES             Location/Qualifiers
     source          1..7110
                     /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             3830..3895
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(3960..3980)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   4373..4513
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4373..4502
                     /label=AAV2 ITR
     rep_origin      4588..5043
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4675..4694)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4885..4906
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5060..5079)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5179..5201
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5239..5257)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5325..5429
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5430..5498,5499..6290)
                     /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(5648..5667)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6461..7049
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6950..6969
                     /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 gggatagatc tagagacctc
     1501 aaaatacatc ctggccaatg ttggggacca gttctgccag ctcgtaatgt ctgagaagga
     1561 ggccatgatg atggtggagc cacaccagaa agtggcatgg aagcgagctg tgcgtggtgt
     1621 acgggagatg tgtgatgtgt gtgaaacaac tctcttcaac atccactggg tttgtcgcaa
     1681 atgtggattt ggggtctgcc ttgactgtta ccggctcagg aaaagccggc cacgcagtga
     1741 gacagaagag atgggtgatg aagaagtttt ctcctggttg aagtgtgcaa agggacagtc
     1801 ccacgaacca gagaatctca tgcccacaca aattattcct ggcacagctc tttacaatat
     1861 tggagacatg gtacatgctg cccggggcaa gtggggaatt aaagcaaact gcccttgtat
     1921 cagtcgacag aacaaatctg tattgagacc tgccgtcacc aatgggatgt cacagcttcc
     1981 tagcataaac cctagtgcct cttctggaaa cgaaactacc ttctctggcg gaggaggacc
     2041 ggcaccagta acaactccag agccggacca tgttcccaaa gccgacagca ctgacatcag
     2101 atctgaagag cctctgaaaa cagacagttc ggcatcaaat agcaatagtg aactgaaagc
     2161 catcaggcct ccttgccctg acacggcccc accctcctcc gccctgcact ggttggcaga
     2221 tttagcaact cagaaggcta aagaagaaac aaaagaagca gggtccctga ggtcggtgct
     2281 caataaagag tctcattcac cctttgggct ggactcgttc aactccactg caaaggtctc
     2341 tccgctgact ccaaagcttt ttaacagtct gttgctgggt cccactgcct ccaacaacaa
     2401 aaccgaaggg tctagccttc gagacctcct tcactccggg ccgggaaaac ttcctcaaac
     2461 ccccttggac acaggcatac cctttccccc ggtcttctct acatcctcag caggagtgaa
     2521 gagcaaggcc agcctaccca actttcttga ccacatcatt gcctcagtgg tagaaaataa
     2581 gaaaacctca gatgcttcaa agcgggcctg caacttgact gatacccaga aggaagtgaa
     2641 ggagatggtg atggggttaa atgtgctaga tccccatact tctcactcct ggctttgtga
     2701 tgggaggctt ctgtgtctcc atgaccccag caacaaaaac aattggaaga tcttccggga
     2761 gtgttggaag caaggtcagc cagtgctggt ttcgggggta cataaaaagc tcaagtctga
     2821 gctctggaag ccagaagcct ttagccagga atttggagac caggatgtag acttggtgaa
     2881 ctgcaggaac tgtgctataa tttccgatgt gaaagttcgg gatttctggg atggtttcga
     2941 gatcatatgc aaacgactac ggtcagaaga tgggcagcca atggtgctca aactcaagga
     3001 ctggcctcct ggggaagatt ttcgagacat gatgccaacc aggtttgaag atctgatgga
     3061 gaaccttcct ctgccagaat ataccaaacg agatggcagg ctcaatctgg cctctaggct
     3121 acctagctac tttgtaaggc ctgatctggg ccccaagatg tacaacgcct atgggttgat
     3181 aacagcagaa gatagaagag ttggtacaac aaatcttcac ttagatgtgt ctgatgctgt
     3241 taatgtgatg gtgtatgttg ggattcccat cggggagggt gctcatgatg aagaggtact
     3301 caagacaatt gacgagggag atgccgatga ggtgacgaag cagaggattc atgatggaaa
     3361 agagaagcca ggtgctttat ggcacatcta tgcagccaag gatgcagaga agatccggga
     3421 gctgctccga aaggttggag aagaacaagg ccaagagaac ccccctgatc atgacccaat
     3481 tcatgaccaa agttggtacc tggaccagac cctccgtaag cgactctatg aggagtatgg
     3541 cgtgcaaggc tgggctattg tacagttcct aggtgatgct gttttcatac ctgctggagc
     3601 cccacaccag gttcacaatc tatacagttg cataaaagta gcagaagact ttgtatctcc
     3661 agaacatgta aagcactgtt tccgcctgac tcaggaattc aggcatctct ctaacactca
     3721 tacaaatcat gaggataaac tgcaggtgaa gaacatcatt taccatgcag tgaaagatgc
     3781 ggttggcacc ctcaaggctc atgaatccaa actggcaagg tcctgtacag actacaaaga
     3841 ccatgacggt gattataaag atcatgacat cgactacaag gatgacgatg acaagtaact
     3901 cgagataatc aacctctgga ttacaaaatt tgtgaaagat tgactggtat tcttaactat
     3961 gttgctcctt ttacgctatg tggatacgct gctttaatgc ctttgtatca tgctattgct
     4021 tcccgtatgg ctttcatttt ctcctccttg tataaatcct ggttagttct tgccacggcg
     4081 gaactcatcg ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt gggcactgac
     4141 aattccgtgg tgtttatttg tgaaatttgt gatgctattg ctttatttgt aaccatctag
     4201 ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa
     4261 acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg agatgtggga
     4321 ggttttttaa agatctggag accggtgtgg cacgtgcgga ccgagcggcc gcaggaaccc
     4381 ctagtgatgg agttggccac tccctctctg cgcgctcgct cgctcactga ggccgggcga
     4441 ccaaaggtcg cccgacgccc gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc
     4501 agctgcctgc aggggcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca
     4561 caccgcatac gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta agcgcggcgg
     4621 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccttagcg cccgctcctt
     4681 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     4741 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     4801 atttgggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     4861 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaact
     4921 ctatctcggg ctattctttt gatttataag ggattttgcc gatttcggtc tattggttaa
     4981 aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta acgtttacaa
     5041 ttttatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagccccgac
     5101 acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca
     5161 gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga
     5221 aacgcgcgag acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa
     5281 taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     5341 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     5401 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     5461 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     5521 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     5581 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     5641 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
     5701 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     5761 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     5821 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     5881 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     5941 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac
     6001 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     6061 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     6121 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     6181 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     6241 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     6301 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     6361 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     6421 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     6481 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     6541 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     6601 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     6661 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     6721 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     6781 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     6841 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     6901 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     6961 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     7021 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     7081 tggccttttg ctggcctttt gctcacatgt
//
