LOCUS       sequence-481012-        6184 bp    DNA     circular SYN 28-JUL-2025
DEFINITION  3' AAVLINK plamid for GRIN2B expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6184)
  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  JOURNAL   Unpublished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 6184)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 28, 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            "Unpublished"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'-GRIN2B-FLAG
FEATURES             Location/Qualifiers
     source          1..6184
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     CDS             288..1319
                     /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     1475..1494
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             2904..2969
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(3034..3054)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   3447..3587
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3447..3576
                     /label=AAV2 ITR
     rep_origin      3662..4117
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3749..3768)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3959..3980
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4134..4153)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4253..4275
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4313..4331)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4399..4503
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4504..4572,4573..5364)
                     /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(4722..4741)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5535..6123
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6024..6043
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgtcgctc tgcgcgaggc ggctccgccg
      181 cggcggaggg atacggcgca ccatatatat atcgcggggc gcagactcgc gctccggcag
      241 tggtgctggg agtgtcgtgg acgccgtgcc gttactcgta cgccaccatg tccaatttac
      301 tgaccgtaca ccaaaatttg cctgcattac cggtcgatgc aacgagtgat gaggttcgca
      361 agaacctgat ggacatgttc agggatcgcc aggcgttttc tgagcatacc tggaaaatgc
      421 ttctgtccgt ttgccggtcg tgggcggcat ggtgcaagtt gaataaccgg aaatggtttc
      481 ccgcagaacc tgaagatgtt cgcgattatc ttctatatct tcaggcgcgc ggtctggcag
      541 taaaaactat ccagcaacat ttgggccagc taaacatgct tcatcgtcgg tccgggctgc
      601 cacgaccaag tgacagcaat gctgtttcac tggttatgcg gcggatccga aaagaaaacg
      661 ttgatgccgg tgaacgtgca aaacaggctc tagcgttcga acgcactgat ttcgaccagg
      721 ttcgttcact catggaaaat agcgatcgct gccaggatat acgtaatctg gcatttctgg
      781 ggattgctta taacaccctg ttacgtatag ccgaaattgc caggatcagg gttaaagata
      841 tctcacgtac tgacggtggg agaatgttaa tccatattgg cagaacgaaa acgctggtta
      901 gcaccgcagg tgtagagaag gcacttagcc tgggggtaac taaactggtc gagcgatgga
      961 tttccgtctc tggtgtagct gatgatccga ataactacct gttttgccgg gtcagaaaaa
     1021 atggtgttgc cgcgccatct gccaccagcc agctatcaac tcgcgccctg gaagggattt
     1081 ttgaagcaac tcatcgattg atttacggcg ctaaggatga ctctggtcag agatacctgg
     1141 cctggtctgg acacagtgcc cgtgtcggag ccgcgcgaga tatggcccgc gctggagttt
     1201 caataccgga gatcatgcaa gctggtggct ggaccaatgt aaatattgta atgaactata
     1261 tccgtaacct ggatagtgaa acaggggcaa tggtgcgcct gctggaagat ggcgattagg
     1321 taagtatcaa ggttacaaga caggtttaag gagaccaata actagtataa cttcgtatag
     1381 gatactttat agcaatttat gaaactgggc ttgtcgagac agagaagact cttgcgtttc
     1441 tgataggcac ctattggtct tactgacatc cactttgcct ttctctccac aggtccatca
     1501 gcaagaagcc cctggacatc ggcctcccct cctccaagca cagccagctc agtgacctgt
     1561 acggcaaatt ctccttcaag agcgaccgct acagtggcca cgacgacttg atccgctccg
     1621 atgtctctga catctcaacc cacaccgtca cctatgggaa catcgagggc aatgccgcca
     1681 agaggcgtaa gcagcaatat aaggacagcc tgaagaagcg gcctgcctcg gccaagtccc
     1741 gcagggagtt tgacgagatc gagctggcct accgtcgccg accgccccgc tcccctgacc
     1801 acaagcgcta cttcagggac aaggaagggc tacgggactt ctacctggac cagttccgaa
     1861 caaaggagaa ctcaccccac tgggagcacg tagacctgac cgacatctac aaggagcgga
     1921 gtgatgactt taagcgcgac tccgtcagcg gaggagggcc ctgtaccaac aggtctcaca
     1981 tcaagcacgg gacgggcgac aaacacggcg tggtcagcgg ggtacctgca ccttgggaga
     2041 agaacctgac caacgtggag tgggaggacc ggtccggggg caacttctgc cgcagctgtc
     2101 cctccaagct gcacaactac tccacgacgg tgacgggtca gaactcgggc aggcaggcgt
     2161 gcatccggtg tgaggcttgc aagaaagcag gcaacctgta tgacatcagt gaggacaact
     2221 ccctgcagga actggaccag ccggctgccc cagtggcggt gacgtcaaac gcctccacca
     2281 ctaagtaccc tcagagcccg actaattcca aggcccagaa gaagaaccgg aacaaactgc
     2341 gccggcagca ctcctacgac accttcgtgg acctgcagaa ggaagaagcc gccctggccc
     2401 cgcgcagcgt aagcctgaaa gacaagggcc gattcatgga tgggagcccc tacgcccaca
     2461 tgtttgagat gtcagctggc gagagcacct ttgccaacaa caagtcctca gtgcccactg
     2521 ccggacatca ccaccacaac aaccccggcg gcgggtacat gctcagcaag tcgctctacc
     2581 ctgaccgggt cacgcaaaac cctttcatcc ccacttttgg ggacgaccag tgcttgctcc
     2641 atggcagcaa atcctacttc ttcaggcagc ccacggtggc gggggcgtcg aaagccaggc
     2701 cggacttccg ggcccttgtc accaacaagc cggtggtctc ggcccttcat ggggccgtgc
     2761 cagcccgttt ccagaaggac atctgtatag ggaaccagtc caacccctgt gtgcctaaca
     2821 acaaaaaccc cagggctttc aatggctcca gcaatgggca tgtttatgag aaactttcta
     2881 gtattgagtc tgatgtctgt acagactaca aagaccatga cggtgattat aaagatcatg
     2941 acatcgacta caaggatgac gatgacaagt aactcgagat aatcaacctc tggattacaa
     3001 aatttgtgaa agattgactg gtattcttaa ctatgttgct ccttttacgc tatgtggata
     3061 cgctgcttta atgcctttgt atcatgctat tgcttcccgt atggctttca ttttctcctc
     3121 cttgtataaa tcctggttag ttcttgccac ggcggaactc atcgccgcct gccttgcccg
     3181 ctgctggaca ggggctcggc tgttgggcac tgacaattcc gtggtgttta tttgtgaaat
     3241 ttgtgatgct attgctttat ttgtaaccat ctagctttat ttgtgaaatt tgtgatgcta
     3301 ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc
     3361 attttatgtt tcaggttcag ggggagatgt gggaggtttt ttaaagatct ggagaccggt
     3421 gtggcacgtg cggaccgagc ggccgcagga acccctagtg atggagttgg ccactccctc
     3481 tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt
     3541 tgcccgggcg gcctcagtga gcgagcgagc gcgcagctgc ctgcaggggc gcctgatgcg
     3601 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atacgtcaaa gcaaccatag
     3661 tacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     3721 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     3781 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     3841 agtgctttac ggcacctcga ccccaaaaaa cttgatttgg gtgatggttc acgtagtggg
     3901 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     3961 ggactcttgt tccaaactgg aacaacactc aaccctatct cgggctattc ttttgattta
     4021 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     4081 aacgcgaatt ttaacaaaat attaacgttt acaattttat ggtgcactct cagtacaatc
     4141 tgctctgatg ccgcatagtt aagccagccc cgacacccgc caacacccgc tgacgcgccc
     4201 tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc
     4261 tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg
     4321 atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc
     4381 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     4441 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
     4501 agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt
     4561 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt
     4621 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc
     4681 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta
     4741 tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac
     4801 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa
     4861 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg
     4921 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc
     4981 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg
     5041 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta
     5101 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg
     5161 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg
     5221 tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc
     5281 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt
     5341 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt
     5401 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     5461 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     5521 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     5581 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     5641 aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag
     5701 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     5761 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     5821 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     5881 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     5941 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     6001 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     6061 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     6121 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     6181 atgt
//
