LOCUS       sequence-476877-        6796 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  3' AAVLINK plasmid for NUP155 expression .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6796)
  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  JOURNAL   Unpublished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 6796)
  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            "Unpublished"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' NUP155
FEATURES             Location/Qualifiers
     source          1..6796
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             2128..2193
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(2258..2278)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   2671..2811
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2671..2800
                     /label=AAV2 ITR
     rep_origin      2886..3341
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2973..2992)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3183..3204
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3358..3377)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3477..3499
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3537..3555)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3623..3727
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3728..3796,3797..4588)
                     /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(3946..3965)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4759..5347
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5248..5267
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   5409..5538
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     CDS             5685..6716
                     /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"
ORIGIN
        1 tgaaactggg cttgtcgaga cagagaagac tcttgcgttt ctgataggca cctattggtc
       61 ttactgacat ccactttgcc tttctctcca cagggtttgc aggaatttct agacagaaac
      121 tcccagtttg caggaggacc attaggaaat ccaaatacta ctgctaaagt gcagcagagg
      181 ctgataggat tcatgcgtcc tgaaaacgga aatccccagc aaatgcaaca ggaactgcag
      241 aggaagtttc atgaggctca actaagtgaa aagatttcac ttcaggcaat tcagcagttg
      301 gttcgaaaat catatcaggc tctggcttta tggaaacttc tttgtgaaca tcaattcact
      361 atcattgtgg cagaacttca gaaggaactt caagagcagc tgaagatcac cacctttaaa
      421 gatcttgtaa tcagggacaa agaactcaca ggggcattaa ttgcttctct tatcaactgc
      481 tacatcagag ataatgccgc tgttgatggc attagtttac atttacagga tatctgccca
      541 cttctatata gcactgatga tgcaatttgt tctaaggcaa atgagcttct ccagcgttcc
      601 cgacaagttc aaaataagac tgaaaaagaa agaatgttaa gggaatcatt aaaggaatat
      661 caaaaaatta gcaatcaagt ggacctttcc aatgtttgtg ctcagtatag acaagtgaga
      721 ttttatgagg gtgtggtgga actttctctt acggctgcag agaaaaaaga tcctcaaggt
      781 cttgggcttc atttctataa acatggagaa ccagaagaag acatagttgg acttcaggcc
      841 ttccaagaaa gattaaacag ttacaaatgc attacagaca cacttcaaga actggtaaat
      901 caaagtaagg ccgctcctca gtctcccagt gtacccaaaa aacctggtcc tccagtgttg
      961 tcatctgatc caaatatgct gagtaatgaa gaagcaggac atcattttga acaaatgctt
     1021 aaattgtcac agcgatccaa ggatgagctc tttagtattg ccctttataa ttggctaata
     1081 caagtcgacc ttgcagataa gctgctacag gttgcttctc catttctgga gccacatcta
     1141 gtccgaatgg ccaaagttga tcaaaacaga gttcgttata tggatttact ctggcggtat
     1201 tacgagaaga acagaagttt cagtaatgct gctcgtgtac tgtccagact ggctgacatg
     1261 catagcacag aaatttcact tcagcagcga ctagagtaca ttgctcgagc cattcttagt
     1321 gccaaaagtt ccactgccat ttcatcaata gctgccgatg gtgaatttct tcatgaatta
     1381 gaagaaaaaa tggaggttgc taggatccaa cttcagatac aggagacact acaaaggcag
     1441 tattcccatc attcttctgt acaggatgca gtttctcagc tggattctga gctgatggac
     1501 ataactaagc tttatgggga atttgctgac ccatttaaac ttgcagagtg caaacttgca
     1561 ataattcatt gtgccggtta ttcagaccct atattggtgc agacactttg gcaagatatc
     1621 atagagaaag aattgagtga cagtgtgaca ttgagctcct cggatagaat gcatgctctt
     1681 agtctcaaga ttgttctcct tggcaaaatt tatgctggca caccacgctt ctttccttta
     1741 gattttattg tacagttttt agaacagcag gtttgtactt tgaactggga tgtgggcttc
     1801 gtaatacaga caatgaatga aattggagta ccattaccta gactactaga agtttatgat
     1861 cagttgttca aatcacggga tccattctgg aacagaatga agaagccact gcaccttttg
     1921 gattgtatac atgtattatt gataagatat gttgagaatc ccagccaagt tttaaattgt
     1981 gaaaggagaa gatttacaaa tctctgcctg gatgctgttt gtggttacct tgttgagctc
     2041 cagtctatga gctcgtcagt agcagtacaa gccatcactg ggaattttaa atctcttcaa
     2101 gctaaattag aacggcttca ttgtacagac tacaaagacc atgacggtga ttataaagat
     2161 catgacatcg actacaagga tgacgatgac aagtaactcg agataatcaa cctctggatt
     2221 acaaaatttg tgaaagattg actggtattc ttaactatgt tgctcctttt acgctatgtg
     2281 gatacgctgc tttaatgcct ttgtatcatg ctattgcttc ccgtatggct ttcattttct
     2341 cctccttgta taaatcctgg ttagttcttg ccacggcgga actcatcgcc gcctgccttg
     2401 cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg tttatttgtg
     2461 aaatttgtga tgctattgct ttatttgtaa ccatctagct ttatttgtga aatttgtgat
     2521 gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc
     2581 attcatttta tgtttcaggt tcagggggag atgtgggagg ttttttaaag atctggagac
     2641 cggtgtggca cgtgcggacc gagcggccgc aggaacccct agtgatggag ttggccactc
     2701 cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc aaaggtcgcc cgacgcccgg
     2761 gctttgcccg ggcggcctca gtgagcgagc gagcgcgcag ctgcctgcag gggcgcctga
     2821 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcatacgt caaagcaacc
     2881 atagtacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
     2941 gaccgctaca cttgccagcg ccttagcgcc cgctcctttc gctttcttcc cttcctttct
     3001 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
     3061 atttagtgct ttacggcacc tcgaccccaa aaaacttgat ttgggtgatg gttcacgtag
     3121 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
     3181 tagtggactc ttgttccaaa ctggaacaac actcaactct atctcgggct attcttttga
     3241 tttataaggg attttgccga tttcggtcta ttggttaaaa aatgagctga tttaacaaaa
     3301 atttaacgcg aattttaaca aaatattaac gtttacaatt ttatggtgca ctctcagtac
     3361 aatctgctct gatgccgcat agttaagcca gccccgacac ccgccaacac ccgctgacgc
     3421 gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg
     3481 gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct
     3541 cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt agacgtcagg
     3601 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     3661 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag
     3721 gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg
     3781 ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt
     3841 gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt
     3901 tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt
     3961 attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa
     4021 tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag
     4081 agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac
     4141 aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac
     4201 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac
     4261 cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac
     4321 tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact
     4381 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg
     4441 tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt
     4501 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat
     4561 aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta
     4621 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     4681 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     4741 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     4801 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     4861 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     4921 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     4981 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     5041 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     5101 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     5161 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     5221 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     5281 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     5341 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     5401 tcacatgtcc tgcaggcagc tgcgcgctcg ctcgctcact gaggccgccc gggcgtcggg
     5461 cgacctttgg tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga gtggccaact
     5521 ccatcactag gggttcctgc ggccgcacgc gtcgctctgc gcgaggcggc tccgccgcgg
     5581 cggagggata cggcgcacca tatatatatc gcggggcgca gactcgcgct ccggcagtgg
     5641 tgctgggagt gtcgtggacg ccgtgccgtt actcgtacgc caccatgtcc aatttactga
     5701 ccgtacacca aaatttgcct gcattaccgg tcgatgcaac gagtgatgag gttcgcaaga
     5761 acctgatgga catgttcagg gatcgccagg cgttttctga gcatacctgg aaaatgcttc
     5821 tgtccgtttg ccggtcgtgg gcggcatggt gcaagttgaa taaccggaaa tggtttcccg
     5881 cagaacctga agatgttcgc gattatcttc tatatcttca ggcgcgcggt ctggcagtaa
     5941 aaaccatcca gcaacatttg ggccagctaa acatgcttca tcgtcggtcc gggctgccac
     6001 gaccaagtga cagcaatgct gtttcactgg ttatgcggcg gatccgaaaa gaaaacgttg
     6061 atgccggtga acgtgcaaaa caggctctag cgttcgaacg cactgatttc gaccaggttc
     6121 gttcactcat ggaaaatagc gatcgctgcc aggatatacg taatctggca tttctgggga
     6181 ttgcttataa caccctgtta cgtatagccg aaattgccag gatcagggtt aaagatatct
     6241 cacgtactga cggtgggaga atgttaatcc atattggcag aacgaaaacg ctggttagca
     6301 ccgcaggtgt agagaaggca cttagcctgg gggtaactaa actggtcgag cgatggattt
     6361 ccgtctctgg tgtagctgat gatccgaata actacctgtt ttgccgggtc agaaaaaatg
     6421 gtgttgccgc gccatctgcc accagccagc tatcaactcg cgccctggaa gggatttttg
     6481 aagcaactca tcgattgatt tacggcgcta aggatgactc tggtcagaga tacctggcct
     6541 ggtctggaca cagtgcccgt gtcggagccg cgcgagatat ggcccgcgct ggagtttcaa
     6601 taccggagat catgcaagct ggtggctgga ccaatgtaaa tattgtaatg aactatatcc
     6661 gtaacctgga tagtgaaaca ggggcaatgg tgcgcctgct ggaagatggc gattaggtaa
     6721 gtatcaaggt tacaagacag gtttaaggag accaataact agtataactt cgtataggat
     6781 actttatagc aattta
//
