LOCUS       sequence-491878-        6856 bp    DNA     circular SYN 04-NOV-2025
DEFINITION  3' AAVLINK plasmid for DIP2C expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6856)
  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  JOURNAL   Unpublished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 6856)
  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            "Unpublished"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'-DIP2C-FLAG
FEATURES             Location/Qualifiers
     source          1..6856
                     /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             3564..3629
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(3706..3726)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   4119..4259
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4119..4248
                     /label=AAV2 ITR
     rep_origin      4334..4789
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4421..4440)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4631..4652
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4806..4825)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4925..4947
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4985..5003)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5071..5175
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5176..5244,5245..6036)
                     /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(5394..5413)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6207..6795
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6696..6715
                     /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 ggcgattgac agtatacatc
     1501 aagttggagt ttattgcctg gccttggtgc cagcaaacac cctccccaaa accccgcttg
     1561 gtgggatcca tttatcagaa acaaaacagc tttttctgga gggctctctg cacccctgca
     1621 atgtcctaat gtgcccccac acctgcgtca caaacttgcc taagcctcga cagaagcagc
     1681 cagaaatcgg ccctgcctct gtgatggtgg ggaacctggt ctctgggaag agaatcgccc
     1741 aggccagtgg cagagacctg ggtcagatcg aagataacga ccaggcacgc aagttcctgt
     1801 tcctctcaga ggtcttgcag tggagagcac agaccacccc ggaccacatc ctctacacgc
     1861 tgctcaactg tcggggtgcg atagcgaact cgctgacctg cgtgcagctg cacaagagag
     1921 ctgagaagat cgccgtgatg ctgatggaga ggggccacct tcaggacggc gaccacgtgg
     1981 ccttggtcta ccccccagga atagacctga tagcagcgtt ttatggttgc ctgtacgcag
     2041 gctgtgtgcc aataaccgtc cgtcccccgc acccacagaa catcgcgacg acgttgccta
     2101 ccgtcaagat gattgtggag gtgagtcgct ctgcctgtct gatgacgaca cagctgatct
     2161 gtaagttgct gcggtccagg gaggcggcgg cggctgtgga cgtcaggacg tggcccctca
     2221 tcctggacac agatgatttg ccaaagaagc ggcctgccca gatctgcaaa ccttgcaacc
     2281 cagacactct tgcatatctc gacttcagcg tgtccacaac tgggatgcta gctggcgtaa
     2341 agatgtctca cgcagccacc agtgccttct gccgttccat taagctgcag tgtgaacttt
     2401 acccctctag agaagtggcc atctgcctgg acccttactg tggactggga tttgtcctct
     2461 ggtgcctctg cagtgtgtat tctgggcacc agtccatcct gatcccgccc tctgagctgg
     2521 aaaccaaccc cgccttgtgg cttcttgccg tgagtcagta caaagtccga gacacgtttt
     2581 gctcctactc cgtgatggag ctgtgcacca aggggctggg ctcgcaaaca gagtccctca
     2641 aggcgcgagg gctggacttg tcccgagtga ggacctgcgt ggttgtggcg gaagagaggc
     2701 ctcggatcgc actcacacag tcgttctcaa agctgtttaa ggacctgggc cttcacccgc
     2761 gggccgtcag cacctcgttc ggttgcaggg tgaacctggc gatttgcttg cagggaacct
     2821 caggacctga cccaaccact gtctacgtgg acatgagagc cctgagacac gacagagtcc
     2881 gcttagtgga aagaggatcc cctcatagtc tgcccctgat ggaatcggga aagatacttc
     2941 caggggttcg gattataatt gccaacccag aaacaaaagg accgctgggg gactcacacc
     3001 ttggagagat ttgggttcac agtgcccaca atgccagcgg ttatttcact atttacggag
     3061 acgaatccct ccagtcagat cacttcaact caagactaag ttttggagac acccagacca
     3121 tctgggcacg cacaggctac ttggggttcc tgcggagaac tgagctcaca gatgcaaatg
     3181 gagagcgcca tgatgccctc tacgtggtag gggcactgga cgaagccatg gagctgcggg
     3241 gcatgcggta ccacccaatc gacattgaga cctcggtcat cagagcccat aaaagcgtta
     3301 cggaatgtgc tgtgtttacc tggacaaatt tgttggtggt tgtggttgag ctggatgggt
     3361 cggaacaaga agccttggac ctggttccct tggtgaccaa cgtggtcctg gaggagcact
     3421 acctgatcgt cggagtggtg gtcgtggtgg acatcggcgt catccccatc aactcccgtg
     3481 gggagaagca gcgcatgcac ctgcgagacg ggtttttggc agaccagcta gaccccatct
     3541 atgtggccta caacatgtgt acagactaca aagaccatga cggtgattat aaagatcatg
     3601 acatcgacta caaggatgac gatgacaagt aagaattctg atgactcgag ataatcaacc
     3661 tctggattac aaaatttgtg aaagattgac tggtattctt aactatgttg ctccttttac
     3721 gctatgtgga tacgctgctt taatgccttt gtatcatgct attgcttccc gtatggcttt
     3781 cattttctcc tccttgtata aatcctggtt agttcttgcc acggcggaac tcatcgccgc
     3841 ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt
     3901 tatttgtgaa atttgtgatg ctattgcttt atttgtaacc atctagcttt atttgtgaaa
     3961 tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa gttaacaaca
     4021 acaattgcat tcattttatg tttcaggttc agggggagat gtgggaggtt ttttaaagat
     4081 ctggagaccg gtgtggcacg tgcggaccga gcggccgcag gaacccctag tgatggagtt
     4141 ggccactccc tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg
     4201 acgcccgggc tttgcccggg cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg
     4261 gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc gcatacgtca
     4321 aagcaaccat agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     4381 cgcagcgtga ccgctacact tgccagcgcc ttagcgcccg ctcctttcgc tttcttccct
     4441 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     4501 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgattt gggtgatggt
     4561 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
     4621 ttctttaata gtggactctt gttccaaact ggaacaacac tcaactctat ctcgggctat
     4681 tcttttgatt tataagggat tttgccgatt tcggtctatt ggttaaaaaa tgagctgatt
     4741 taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt atggtgcact
     4801 ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc
     4861 gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc
     4921 gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgagacga
     4981 aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag
     5041 acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
     5101 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     5161 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     5221 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     5281 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     5341 gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt
     5401 ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat
     5461 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     5521 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     5581 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     5641 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     5701 cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa
     5761 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     5821 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc
     5881 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt
     5941 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     6001 gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat
     6061 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     6121 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     6181 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     6241 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     6301 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta
     6361 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     6421 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     6481 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     6541 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     6601 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     6661 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     6721 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     6781 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     6841 ccttttgctc acatgt
//
