LOCUS       sequence-491870-        5661 bp    DNA     circular SYN 04-NOV-2025
DEFINITION  3' AAVLINK plasmid for ARHGAP5 expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5661)
  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  JOURNAL   Unpublished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 5661)
  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            "Unpublished"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'-ARHGAP5-FLAG
FEATURES             Location/Qualifiers
     source          1..5661
                     /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"
     primer_bind     1464..1483
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             2369..2434
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(2511..2531)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   2924..3064
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2924..3053
                     /label=AAV2 ITR
     rep_origin      3139..3594
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3226..3245)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3436..3457
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3611..3630)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3730..3752
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3790..3808)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3876..3980
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3981..4049,4050..4841)
                     /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(4199..4218)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5012..5600
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5501..5520
                     /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 ggtggtattg ataatcctgc
     1501 aatcacttct gaccaggagt tagatgataa gaagatgaag aagaaaaccc acaaagtgaa
     1561 agaagataaa aagcagaaaa agaaaactaa gaacttcaat ccaccaacac gtagaaattg
     1621 ggaaagtaat tactttggga tgcccctcca ggatctggtt acagctgaga agcccatacc
     1681 actatttgtt gagaaatgtg tggaatttat tgaagataca gggttatgta ccgaaggact
     1741 ctaccgtgtc agcgggaata aaactgacca agacaatatt caaaagcagt ttgatcaaga
     1801 tcataatatc aatctagtgt caatggaagt aacagtaaat gctgtagctg gagcccttaa
     1861 agctttcttt gcagatctgc cagatccttt aattccatat tctcttcatc cagaactatt
     1921 ggaagcagca aaaatcccgg ataaaacaga acgtcttcat gccttgaaag aaattgttaa
     1981 gaaatttcat cctgtaaact atgatgtatt cagatacgtg ataacacatc taaacagggt
     2041 tagtcagcaa cataaaatca acctaatgac agcagacaac ttatccatct gtttttggcc
     2101 aaccttgatg agacctgatt ttgaaaatcg agagtttctg tctactacta agattcatca
     2161 atctgttgtt gaaacattca ttcagcagtg tcagtttttc ttttacaatg gagaaattgt
     2221 agaaacgaca aacattgtgg ctcctccacc accttcaaac ccaggacagt tggtggaacc
     2281 aatggtgcca cttcagttgc cgccaccatt gcaacctcag ctgatacaac cacaattaca
     2341 aacggatcct cttggtatta tatgtacaga ctacaaagac catgacggtg attataaaga
     2401 tcatgacatc gactacaagg atgacgatga caagtaagaa ttctgatgac tcgagataat
     2461 caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta tgttgctcct
     2521 tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc ttcccgtatg
     2581 gctttcattt tctcctcctt gtataaatcc tggttagttc ttgccacggc ggaactcatc
     2641 gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt tgggcactga caattccgtg
     2701 gtgtttattt gtgaaatttg tgatgctatt gctttatttg taaccatcta gctttatttg
     2761 tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa
     2821 caacaacaat tgcattcatt ttatgtttca ggttcagggg gagatgtggg aggtttttta
     2881 aagatctgga gaccggtgtg gcacgtgcgg accgagcggc cgcaggaacc cctagtgatg
     2941 gagttggcca ctccctctct gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc
     3001 gcccgacgcc cgggctttgc ccgggcggcc tcagtgagcg agcgagcgcg cagctgcctg
     3061 caggggcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata
     3121 cgtcaaagca accatagtac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     3181 ttacgcgcag cgtgaccgct acacttgcca gcgccttagc gcccgctcct ttcgctttct
     3241 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     3301 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gatttgggtg
     3361 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     3421 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac tctatctcgg
     3481 gctattcttt tgatttataa gggattttgc cgatttcggt ctattggtta aaaaatgagc
     3541 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgtttaca attttatggt
     3601 gcactctcag tacaatctgc tctgatgccg catagttaag ccagccccga cacccgccaa
     3661 cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg
     3721 tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga
     3781 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
     3841 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     3901 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     3961 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     4021 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     4081 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     4141 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     4201 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     4261 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     4321 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     4381 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     4441 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     4501 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
     4561 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     4621 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     4681 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     4741 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     4801 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     4861 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     4921 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     4981 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     5041 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     5101 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     5161 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     5221 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     5281 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     5341 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     5401 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     5461 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     5521 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     5581 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     5641 gctggccttt tgctcacatg t
//
