LOCUS       sequence-489643-        6766 bp    DNA     circular SYN 17-OCT-2025
DEFINITION  3' AAVLINK plasmid for BRD4 expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6766)
  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  JOURNAL   Unpublished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 6766)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 17, 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            "Unpublished"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' BRD4
FEATURES             Location/Qualifiers
     source          1..6766
                     /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             1605..1622
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             3486..3551
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(3616..3636)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   4029..4169
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4029..4158
                     /label=AAV2 ITR
     rep_origin      4244..4699
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4331..4350)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4541..4562
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4716..4735)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4835..4857
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4895..4913)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4981..5085
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5086..5154,5155..5946)
                     /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(5304..5323)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6117..6705
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6606..6625
                     /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 gggcttctcg tcctcagagt
     1501 cggagagctc cagtgagtcc agctcctctg acagcgaaga ctccgaaaca gagatggctc
     1561 cgaagtcaaa aaagaagggg caccccggga gggagcagaa gaagcaccat catcaccacc
     1621 atcagcagat gcagcaggcc ccggctcctg tgccccagca gccgcccccg cctccccagc
     1681 agcccccacc gcctccacct ccgcagcagc aacagcagcc gccacccccg cctcccccac
     1741 cctccatgcc gcagcaggca gccccggcga tgaagtcctc gcccccaccc ttcattgcca
     1801 cccaggtgcc cgtcctggag ccccagctcc caggcagcgt ctttgacccc atcggccact
     1861 tcacccagcc catcctgcac ctgccgcagc ctgagctgcc ccctcacctg ccccagccgc
     1921 ctgagcacag cactccaccc catctcaacc agcacgcagt ggtctctcct ccagctttgc
     1981 acaacgcact accccagcag ccatcacggc ccagcaaccg agccgctgcc ctgcctccca
     2041 agcccgcccg gcccccagcc gtgtcaccag ccttgaccca aacacccctg ctcccacagc
     2101 cccccatggc ccaacccccc caagtgctgc tggaggatga agagccacct gccccacccc
     2161 tcacctccat gcagatgcag ctgtacctgc agcagctgca gaaggtgcag ccccctacgc
     2221 cgctactccc ttccgtgaag gtgcagtccc agcccccacc ccccctgccg cccccacccc
     2281 acccctctgt gcagcagcag ctgcagcagc agccgccacc acccccacca ccccagcccc
     2341 agcctccacc ccagcagcag catcagcccc ctccacggcc cgtgcacttg cagcccatgc
     2401 agttttccac ccacatccaa cagcccccgc caccccaggg ccagcagccc ccccatccgc
     2461 ccccaggcca gcagccaccc ccgccgcagc ctgccaagcc tcagcaagtc atccagcacc
     2521 accattcacc ccggcaccac aagtcggacc cctactcaac cggtcacctc cgcgaagccc
     2581 cctccccgct tatgatacat tccccccaga tgtcacagtt ccagagcctg acccaccagt
     2641 ctccacccca gcaaaacgtc cagcctaaga aacaggagct gcgtgctgcc tccgtggtcc
     2701 agccccagcc cctcgtggtg gtgaaggagg agaagatcca ctcacccatc atccgcagcg
     2761 agcccttcag cccctcgctg cggccggagc cccccaagca cccggagagc atcaaggccc
     2821 ccgtccacct gccccagcgg ccggaaatga agcctgtgga tgtcgggagg cctgtgatcc
     2881 ggcccccaga gcagaacgca ccgccaccag gggcccctga caaggacaaa cagaaacagg
     2941 agccgaagac tccagttgcg cccaaaaagg acctgaaaat caagaacatg ggctcctggg
     3001 ccagcctagt gcagaagcat ccgaccaccc cctcctccac agccaagtca tccagcgaca
     3061 gcttcgagca gttccgccgc gccgctcggg agaaagagga gcgtgagaag gccctgaagg
     3121 ctcaggccga gcacgctgag aaggagaagg agcggctgcg gcaggagcgc atgaggagcc
     3181 gagaggacga ggatgcgctg gagcaggccc ggcgggccca tgaggaggca cgtcggcgcc
     3241 aggagcagca gcagcagcag cgccaggagc aacagcagca gcagcaacag caagcagctg
     3301 cggtggctgc cgccgccacc ccacaggccc agagctccca gccccagtcc atgctggacc
     3361 agcagaggga gttggcccgg aagcgggagc aggagcgaag acgccgggaa gccatggcag
     3421 ctaccattga catgaatttc cagagtgatc tattgtcaat atttgaagaa aatcttttct
     3481 gtacagacta caaagaccat gacggtgatt ataaagatca tgacatcgac tacaaggatg
     3541 acgatgacaa gtaactcgag ataatcaacc tctggattac aaaatttgtg aaagattgac
     3601 tggtattctt aactatgttg ctccttttac gctatgtgga tacgctgctt taatgccttt
     3661 gtatcatgct attgcttccc gtatggcttt cattttctcc tccttgtata aatcctggtt
     3721 agttcttgcc acggcggaac tcatcgccgc ctgccttgcc cgctgctgga caggggctcg
     3781 gctgttgggc actgacaatt ccgtggtgtt tatttgtgaa atttgtgatg ctattgcttt
     3841 atttgtaacc atctagcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca
     3901 ttataagctg caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc
     3961 agggggagat gtgggaggtt ttttaaagat ctggagaccg gtgtggcacg tgcggaccga
     4021 gcggccgcag gaacccctag tgatggagtt ggccactccc tctctgcgcg ctcgctcgct
     4081 cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg cggcctcagt
     4141 gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg cggtattttc tccttacgca
     4201 tctgtgcggt atttcacacc gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc
     4261 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     4321 ttagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     4381 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     4441 gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     4501 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     4561 ggaacaacac tcaactctat ctcgggctat tcttttgatt tataagggat tttgccgatt
     4621 tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     4681 atattaacgt ttacaatttt atggtgcact ctcagtacaa tctgctctga tgccgcatag
     4741 ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
     4801 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
     4861 tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag
     4921 gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg
     4981 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     5041 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
     5101 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
     5161 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
     5221 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
     5281 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg
     5341 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
     5401 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     5461 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     5521 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     5581 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
     5641 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     5701 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     5761 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
     5821 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     5881 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     5941 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     6001 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     6061 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     6121 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     6181 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     6241 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     6301 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     6361 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     6421 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     6481 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     6541 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     6601 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     6661 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     6721 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgt
//
