LOCUS       sequence-491857-        7092 bp    DNA     circular SYN 04-NOV-2025
DEFINITION  3' AAVLINK plasmid for ASTN2 expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7092)
  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  JOURNAL   Unpublished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 7092)
  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            "Unpublished"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'-ASTN2-HA
FEATURES             Location/Qualifiers
     source          1..7092
                     /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             3791..3817
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3821..3847
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3851..3877
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(3942..3962)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   4355..4495
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4355..4484
                     /label=AAV2 ITR
     rep_origin      4570..5025
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4657..4676)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4867..4888
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5042..5061)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5161..5183
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5221..5239)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5307..5411
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5412..5480,5481..6272)
                     /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(5630..5649)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6443..7031
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6932..6951
                     /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 ggggagcaag cccctgaaaa
     1501 gattctcagg tctactttca gcttgggcca aggcctctgg cttcctgtca gcaaaagctt
     1561 tgtggttccg cctgtggagc tgtccatcaa ccccctggcc agctgcaaga ccgatgtgct
     1621 cgtcacggaa gaccctgcag atgtcaggga agaagcgatg ctgtccacat actttgaaac
     1681 catcaatgac ctgctgtctt ccttcgggcc agttcgtgac tgctctcgga acaatggggg
     1741 ctgcactcgc aacttcaagt gtgtgtctga ccggcaggtg gattcctcgg gatgtgtgtg
     1801 ccctgaggag ctgaaaccca tgaaggatgg ctctggctgc tacgaccact ccaaaggcat
     1861 tgactgctct gatggcttta atggcggctg tgagcagctg tgcctgcagc agacgctgcc
     1921 cctgccctac gatgccactt cgagcaccat cttcatgttc tgcggttgcg tggaggagta
     1981 caaactggct cctgatggaa aatcctgctt aatgctctca gatgtctgcg agggccccaa
     2041 gtgcctcaaa cctgactcca aattcaatga taccctcttt ggagagatgc tacatggtta
     2101 caacaaccgg acccagcatg tgaaccaagg ccaagtcttc cagatgacct ttagggagaa
     2161 caacttcatc aaggactttc cccagctggc cgatgggctg ttggtgatcc cgctgccggt
     2221 ggaggagcag tgccgggggg tcctctccga gccccttccg gacctccaac tgctcactgg
     2281 agatatcagg tatgatgagg ccatgggtta ccccatggtg cagcagtggc gggtccggag
     2341 caacctctac cgtgtgaagc tcagcaccat caccctcgca gcaggcttca ctaatgttct
     2401 caagatcctg accaaggaga gcagtcggga ggagctgctg tccttcatcc agcactatgg
     2461 ctcccactac atcgcagagg ccctctatgg ctcagagctc acctgcatca tccactttcc
     2521 cagcaagaag gtccagcagc agctgtggct ccagtatcag aaagagacca cagagctggg
     2581 cagcaagaag gagctcaagt ccatgccctt catcacctac ctctcaggtt tgctgacagc
     2641 ccagatgctg tcagatgacc agctcatttc aggtgtggag attcgctgtg aggagaaggg
     2701 gcgctgtcca tctacctgtc acctttgccg ccggccaggc aaggagcagc tgagccccac
     2761 accagtgctg ctggaaatca accgtgtggt gccactttat accctcatcc aagacaatgg
     2821 cacaaaggag gccttcaaga gtgcactgat gagttcctac tggtgctcag ggaaagggga
     2881 tgtgatcgat gactggtgca ggtgtgacct cagcgccttt gatgccaatg ggctccccaa
     2941 ctgcagcccc cttctgcagc cggtgctgcg gctgtcccca acagtggagc cctccagtac
     3001 tgtggtctcc ttggagtggg tggatgttca gccagctatt gggaccaagg tctccgacta
     3061 tattctgcag cataagaaag tggatgaata cacagacact gacctgtaca caggagaatt
     3121 cctgagtttt gctgatgact tactctctgg cctgggcaca tcttgtgtag cagctggtcg
     3181 aagccatgga gaggtccctg aagtcagtat ctactcagtc atcttcaagt gtctggagcc
     3241 cgacggtctc tacaagttca ctctgtatgc tgtggataca cgagggaggc actcagagct
     3301 aagcacggtg accctgagga cggcctgtcc actggtagat gacaacaagg cagaagaaat
     3361 agctgacaag atctacaatc tgtacaatgg gtacacaagt ggaaaggagc agcagatggc
     3421 ctacaacaca ctgatggagg tctcagcctc gatgctgttc cgagtccagc accactacaa
     3481 ctctcactat gaaaagtttg gcgacttcgt ctggagaagt gaggatgagc tggggcccag
     3541 gaaggcccac ctgattctac ggcgactgga gagggtgagt agccactgct ccagcctcct
     3601 gcggagtgcc tacatccaga gccgcgtgga aacagtgccc tatcttttct gccgcagcga
     3661 ggaggtccgg cctgcaggca tggtgtggta tagcatcctc aaggacacca aaatcacgtg
     3721 tgaggagaag atggtgtcaa tggcccgaaa cacgtacggg gagtccaagg gccggggagg
     3781 tggatctggt tacccgtatg atgttccgga ttacgctggc tatccctacg acgtgcccga
     3841 ctatgccggg tacccctatg acgtcccaga ctacgcataa ctcgagataa tcaacctctg
     3901 gattacaaaa tttgtgaaag attgactggt attcttaact atgttgctcc ttttacgcta
     3961 tgtggatacg ctgctttaat gcctttgtat catgctattg cttcccgtat ggctttcatt
     4021 ttctcctcct tgtataaatc ctggttagtt cttgccacgg cggaactcat cgccgcctgc
     4081 cttgcccgct gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgtttatt
     4141 tgtgaaattt gtgatgctat tgctttattt gtaaccatct agctttattt gtgaaatttg
     4201 tgatgctatt gctttatttg taaccattat aagctgcaat aaacaagtta acaacaacaa
     4261 ttgcattcat tttatgtttc aggttcaggg ggagatgtgg gaggtttttt aaagatctgg
     4321 agaccggtgt ggcacgtgcg gaccgagcgg ccgcaggaac ccctagtgat ggagttggcc
     4381 actccctctc tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc
     4441 ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc gcagctgcct gcaggggcgc
     4501 ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat acgtcaaagc
     4561 aaccatagta cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     4621 gcgtgaccgc tacacttgcc agcgccttag cgcccgctcc tttcgctttc ttcccttcct
     4681 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     4741 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgatttgggt gatggttcac
     4801 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct
     4861 ttaatagtgg actcttgttc caaactggaa caacactcaa ctctatctcg ggctattctt
     4921 ttgatttata agggattttg ccgatttcgg tctattggtt aaaaaatgag ctgatttaac
     4981 aaaaatttaa cgcgaatttt aacaaaatat taacgtttac aattttatgg tgcactctca
     5041 gtacaatctg ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg
     5101 acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct
     5161 ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg
     5221 gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt
     5281 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     5341 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     5401 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     5461 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     5521 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     5581 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     5641 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     5701 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     5761 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     5821 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     5881 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     5941 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     6001 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     6061 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     6121 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     6181 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     6241 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     6301 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     6361 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     6421 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     6481 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     6541 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     6601 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     6661 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     6721 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     6781 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     6841 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     6901 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     6961 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     7021 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     7081 ttgctcacat gt
//
