LOCUS       sequence-491444-        7205 bp    DNA     circular SYN 03-NOV-2025
DEFINITION  3' AAVLINK plasmid for PRR14L expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7205)
  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  JOURNAL   Unpublished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 7205)
  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            "Unpublished"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'-PRR14L-FLAG
FEATURES             Location/Qualifiers
     source          1..7205
                     /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             3913..3978
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(4055..4075)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   4468..4608
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4468..4597
                     /label=AAV2 ITR
     rep_origin      4683..5138
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4770..4789)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4980..5001
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5155..5174)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5274..5296
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5334..5352)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5420..5524
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5525..5593,5594..6385)
                     /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(5743..5762)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6556..7144
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7045..7064
                     /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 ggggacgact gggttactta
     1501 actgttgggg agcaatctga ggagttggtt accagagaaa ctggcgatgg cgatcccgtg
     1561 agcaacatct ctcagaccca ttttaaatgc cgggggatac ttaatcatgc tgaaaaacag
     1621 cagagccctg aggttttgga ctacatgttg cagaaagaag agaaatatat acgtcaacaa
     1681 aaagcacata cgatatcgca acagtgcata tcatctagtc tgttgttaga tgatgcacaa
     1741 aatcagaacc aaccgaaggc tgacaaagat gagtccacca tgatcaatga aatcacccta
     1801 gcaaagctgg ccaaggacag cattgtggca cagactcaga agttagaaga ccagaaggag
     1861 gaaaggttac atcatccatt aaggaaggac actgagtcat gcacaagtcc ttgccttctt
     1921 ggtgcccctc ggaaagcaca agacccctcc tctgctgggt gtgatcaaat acatggtgcc
     1981 tttgcgaaga aaggagttct tcccttaaag aagcagcccc atcgaacttg taagaaagtt
     2041 tcctatcagg agcaaatcat tgttgggaga aaaataggta aaatcagaag ttctgccttt
     2101 ttaaagagtt cttccaatcc catccccaca aaagcgcaca gacttctcag tttgtgtact
     2161 ctgtctgcac ctacacgatt agaacctgaa acagcaccta ccaagagctt ggttagtcac
     2221 ataccaaagc agatgtctac accgtgccat cccttgagga gcctgaattt taggaagact
     2281 accaaagaat cagccttact aaacaagctg tccatccttg cctccaaact ggccccagcc
     2341 atgaagactc agaagctaag ataccgacgg tgttcctctg aacttcttcc aatggctaaa
     2401 agctacaagc gcctcagata taaaagactc ctggacggat tttcatccag cacagagcag
     2461 ctgaatccat atttggcagc tagtggatgg gataagaggc ctaacagtaa gcccatggca
     2521 ctttattctc tcgaatccat caagatgacc ttcatagatt tgagcaacaa gatgccgtcc
     2581 ctgctgtttg gttctgaaat cttcccagta tcctttcatg tgaaatcatc cagctcagat
     2641 tgcacgactg agtcctcaag gacttttcct gagcactgtg ctccggcaag gcttgcctta
     2701 ggagaggccc tccagtgccc gtctcaacct cccaagtgga ccttttcttt cttcttgtcc
     2761 cacggttgcc ctgggatggc aacattcagg gaagacactg gcgtccatag tcagacccac
     2821 actcaggctc ctccccagcc tccagctcct ctccaagact atggaggcac tgccatagtc
     2881 cagaccagag cagactgctc tgtccttggc cttcacacac ttctagcact ttgttcccca
     2941 ggatgttacc gaatctggac aaaaaaacgg agcttctcca gccacatgcc taccatgcag
     3001 aggctcttca tgacccagtt tacacagggc ctgaaagggt tacggtctcc agcctccata
     3061 gcagacaagg tcttctgttc tctgccctac tcggtgggca gagtcctatc catttggagc
     3121 cagcatggtc cttctgtctg ctccttcgaa atctcttctc ttcattcccc tcactgcaag
     3181 cggcaaccaa gtctgggcac cacaagcagc cacaccatgt taccatatgt gcctcttcca
     3241 ggcatggaag ctacatataa caccagcggc agtcagacga ggctggagcc tccattccct
     3301 gccttggtac caaagtcttg cttggtagca gaatcagctg tcagcaagct cctgctttca
     3361 gcctctgagt tccaggttcg tggattggat gagctggatg gtgtgaaagc agcatgcccc
     3421 tgcccacaga gcagcccccc agaacagaaa gaggctgagc cagagaagag gccaaagaaa
     3481 gtctcacaga ttcgcatccg gaaaaccatt cctaggccag atcctaatct tacccccatg
     3541 ggccttcctc gacccaaaag gttaaagaag aaggagttta gtttagaaga gatatatacc
     3601 aacaagaatt ataaatctcc tcctgcaaac aggtgtttag agaccatctt tgaggaaccc
     3661 aaggaacgaa atggtacact aatctcaatc agccaacaga agaggaagcg agttctagaa
     3721 tttcaggatt ttacagtccc gcgaaagagg agagctcgag gcaaagtcaa ggtggcaggc
     3781 agctttacca gggcccagaa ggcagctgtg cagagtcgag agctggatgc tcttttgata
     3841 cagaaactaa tggaactgga gaccttcttt gccaaggaag aggagcagga acaatcatca
     3901 ggctgttgta cagactacaa agaccatgac ggtgattata aagatcatga catcgactac
     3961 aaggatgacg atgacaagta agaattctga tgactcgaga taatcaacct ctggattaca
     4021 aaatttgtga aagattgact ggtattctta actatgttgc tccttttacg ctatgtggat
     4081 acgctgcttt aatgcctttg tatcatgcta ttgcttcccg tatggctttc attttctcct
     4141 ccttgtataa atcctggtta gttcttgcca cggcggaact catcgccgcc tgccttgccc
     4201 gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttt atttgtgaaa
     4261 tttgtgatgc tattgcttta tttgtaacca tctagcttta tttgtgaaat ttgtgatgct
     4321 attgctttat ttgtaaccat tataagctgc aataaacaag ttaacaacaa caattgcatt
     4381 cattttatgt ttcaggttca gggggagatg tgggaggttt tttaaagatc tggagaccgg
     4441 tgtggcacgt gcggaccgag cggccgcagg aacccctagt gatggagttg gccactccct
     4501 ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct
     4561 ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg cctgcagggg cgcctgatgc
     4621 ggtattttct ccttacgcat ctgtgcggta tttcacaccg catacgtcaa agcaaccata
     4681 gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     4741 cgctacactt gccagcgcct tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     4801 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
     4861 tagtgcttta cggcacctcg accccaaaaa acttgatttg ggtgatggtt cacgtagtgg
     4921 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     4981 tggactcttg ttccaaactg gaacaacact caactctatc tcgggctatt cttttgattt
     5041 ataagggatt ttgccgattt cggtctattg gttaaaaaat gagctgattt aacaaaaatt
     5101 taacgcgaat tttaacaaaa tattaacgtt tacaatttta tggtgcactc tcagtacaat
     5161 ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg ctgacgcgcc
     5221 ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag
     5281 ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa agggcctcgt
     5341 gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg
     5401 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     5461 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     5521 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     5581 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     5641 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     5701 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     5761 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     5821 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     5881 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     5941 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     6001 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     6061 gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct
     6121 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
     6181 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
     6241 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
     6301 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     6361 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat
     6421 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
     6481 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     6541 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     6601 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     6661 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
     6721 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     6781 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     6841 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     6901 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     6961 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     7021 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     7081 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     7141 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     7201 catgt
//
