LOCUS       sequence-491865-        6660 bp    DNA     circular SYN 04-NOV-2025
DEFINITION  3' AAVLINK plasmid for CECR2 expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6660)
  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  JOURNAL   Unpublished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 6660)
  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            "Unpublished"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'-CECR2-FLAG
FEATURES             Location/Qualifiers
     source          1..6660
                     /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             3368..3433
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(3510..3530)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   3923..4063
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3923..4052
                     /label=AAV2 ITR
     rep_origin      4138..4593
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4225..4244)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4435..4456
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4610..4629)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4729..4751
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4789..4807)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4875..4979
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4980..5048,5049..5840)
                     /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(5198..5217)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6011..6599
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6500..6519
                     /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 gggtacatgc gaccgccctg
     1501 caagtctgcc ggacatcggt tacagccacc tccagtgcca gcacccagtt ctttgtttgg
     1561 agcacctgcc caggctcttc ggggggtgca gggaggggac tccatgatgg acagcccaga
     1621 gatgattgcg atgcagcagc tctcctcccg cgtctgcccc ccaggtgtgc cttaccaccc
     1681 ccaccagcct gcacaccccc gtttacctgg cccttttccg caggtagctc acccaatgtc
     1741 agtcactgtg tcagccccca agcctgccct gggcaaccct gggagggcac cggagaacag
     1801 tgaagcacaa gagcctgaga atgaccaagc agagccgttg cctggccttg aagagaaacc
     1861 accaggtgtt ggtacttcag agggggtcta cctcacacaa ctacctcacc ccacacctcc
     1921 cctgcagact gactgcacca ggcagagctc accacaagaa agggaaacag tgggcccgga
     1981 gctcaaaagc agctcctccg aatctgcgga caactgtaaa gcaatgaagg gcaagaatcc
     2041 ctggccctcg gatagcagct accccggccc agccgcccaa gggtgcgtga gagacctctc
     2101 cacggtggca gacaggggcg ctctatccga gaacggagtc attggggaag catctccttg
     2161 tggatcggag gggaagggcc ttggtagcag tggttccgaa aagctgctct gccccagagg
     2221 cagaacgttg caggaaacca tgccatgcac gggacagaac gcagcgacac cgcccagcac
     2281 agaccccggt ttgacgggag gcactgtgag ccagtttccc ccgctgtata tgcctggcct
     2341 agagtacccg aattcagctg cccattacca catcagtcca ggcctgcagg gtgtgggccc
     2401 tgtgatggga gggaagtccc cagcatccca tccccagcat tttcccccaa ggggctttca
     2461 gtctaaccac ccacattctg gaggctttcc ccggtatcgc cccccacaag gaatgaggta
     2521 ttcctaccac ccaccgccac agccttccta ccaccactat cagcgaactc cttactatgc
     2581 ctgtccacag agcttttctg actggcagag acctctccat ccccagggaa gcccaagcgg
     2641 acccccagcc agtcagcctc ccccaccaag gtccctcttc tcagataaga atgccatggc
     2701 cagtctgcaa ggctgtgaga cactgaatgc tgccttaact tctccaaccc gtatggatgc
     2761 agtggctgct aaagtcccaa atgacgggca gaatcctggt ccagaggaag agaagctgga
     2821 tgaatctatg gagaggccag agagtcccaa agaattttta gacctggaca accataacgc
     2881 agctaccaag cggcagagct cgttgtcagc cagcgagtat ctctatggaa ctcctccgcc
     2941 tctgagttca ggaatgggat ttggttcatc tgcatttcca ccccacagtg tgatgctgca
     3001 gacggggcct ccctataccc ctcagcggcc ggccagtcac tttcagccca gggcttactc
     3061 ttcccctgtg gctgccctcc cacctcacca cccaggggcc acccagccca acggcctctc
     3121 tcaggagggt cccatctatc gctgccagga agaaggcctg ggtcactttc aagctgtgat
     3181 gatggaacaa attggcacta gaagtggaat aagaggacct ttccaggaaa tgtacagacc
     3241 atcaggaatg cagatgcacc cggtccagtc gcaggcctcg ttcccaaaga cccccacagc
     3301 agcaacatca caggaggagg tgccgcctca taagcctcca acacttcccc tggatcagag
     3361 ctgtacagac tacaaagacc atgacggtga ttataaagat catgacatcg actacaagga
     3421 tgacgatgac aagtaagaat tctgatgact cgagataatc aacctctgga ttacaaaatt
     3481 tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct
     3541 gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg
     3601 tataaatcct ggttagttct tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc
     3661 tggacagggg ctcggctgtt gggcactgac aattccgtgg tgtttatttg tgaaatttgt
     3721 gatgctattg ctttatttgt aaccatctag ctttatttgt gaaatttgtg atgctattgc
     3781 tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt gcattcattt
     3841 tatgtttcag gttcaggggg agatgtggga ggttttttaa agatctggag accggtgtgg
     3901 cacgtgcgga ccgagcggcc gcaggaaccc ctagtgatgg agttggccac tccctctctg
     3961 cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc
     4021 cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct gatgcggtat
     4081 tttctcctta cgcatctgtg cggtatttca caccgcatac gtcaaagcaa ccatagtacg
     4141 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     4201 cacttgccag cgccttagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     4261 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     4321 ctttacggca cctcgacccc aaaaaacttg atttgggtga tggttcacgt agtgggccat
     4381 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     4441 tcttgttcca aactggaaca acactcaact ctatctcggg ctattctttt gatttataag
     4501 ggattttgcc gatttcggtc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     4561 cgaattttaa caaaatatta acgtttacaa ttttatggtg cactctcagt acaatctgct
     4621 ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac
     4681 gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca
     4741 tgtgtcagag gttttcaccg tcatcaccgg aacgcgcgag acgaaagggc ctcgtgatac
     4801 gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt
     4861 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     4921 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
     4981 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     5041 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
     5101 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     5161 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     5221 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     5281 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     5341 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     5401 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     5461 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     5521 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     5581 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     5641 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     5701 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     5761 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     5821 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     5881 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     5941 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     6001 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     6061 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     6121 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
     6181 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     6241 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     6301 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     6361 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     6421 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     6481 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     6541 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     6601 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
//
