LOCUS       sequence-491388-        6173 bp    DNA     circular SYN 03-NOV-2025
DEFINITION  3' AAVLINK plasmid for GRIN2B expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6173)
  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  JOURNAL   Unpublished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 6173)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 3, 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            "Unpublished"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..6173
                     /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             2893..2958
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(3023..3043)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   3436..3576
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3436..3565
                     /label=AAV2 ITR
     rep_origin      3651..4106
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3738..3757)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3948..3969
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4123..4142)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4242..4264
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4302..4320)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4388..4492
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4493..4561,4562..5353)
                     /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(4711..4730)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5524..6112
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6013..6032
                     /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 ggtccatcag caagaagccc
     1501 ctggacatcg gcctcccctc ctccaagcac agccagctca gtgacctgta cggcaaattc
     1561 tccttcaaga gcgaccgcta cagtggccac gacgacttga tccgctccga tgtctctgac
     1621 atctcaaccc acaccgtcac ctatgggaac atcgagggca atgccgccaa gaggcgtaag
     1681 cagcaatata aggacagcct gaagaagcgg cctgcctcgg ccaagtcccg cagggagttt
     1741 gacgagatcg agctggccta ccgtcgccga ccgccccgct cccctgacca caagcgctac
     1801 ttcagggaca aggaagggct acgggacttc tacctggacc agttccgaac aaaggagaac
     1861 tcaccccact gggagcacgt agacctgacc gacatctaca aggagcggag tgatgacttt
     1921 aagcgcgact ccgtcagcgg aggagggccc tgtaccaaca ggtctcacat caagcacggg
     1981 acgggcgaca aacacggcgt ggtcagcggg gtacctgcac cttgggagaa gaacctgacc
     2041 aacgtggagt gggaggaccg gtccgggggc aacttctgcc gcagctgtcc ctccaagctg
     2101 cacaactact ccacgacggt gacgggtcag aactcgggca ggcaggcgtg catccggtgt
     2161 gaggcttgca agaaagcagg caacctgtat gacatcagtg aggacaactc cctgcaggaa
     2221 ctggaccagc cggctgcccc agtggcggtg acgtcaaacg cctccaccac taagtaccct
     2281 cagagcccga ctaattccaa ggcccagaag aagaaccgga acaaactgcg ccggcagcac
     2341 tcctacgaca ccttcgtgga cctgcagaag gaagaagccg ccctggcccc gcgcagcgta
     2401 agcctgaaag acaagggccg attcatggat gggagcccct acgcccacat gtttgagatg
     2461 tcagctggcg agagcacctt tgccaacaac aagtcctcag tgcccactgc cggacatcac
     2521 caccacaaca accccggcgg cgggtacatg ctcagcaagt cgctctaccc tgaccgggtc
     2581 acgcaaaacc ctttcatccc cacttttggg gacgaccagt gcttgctcca tggcagcaaa
     2641 tcctacttct tcaggcagcc cacggtggcg ggggcgtcga aagccaggcc ggacttccgg
     2701 gcccttgtca ccaacaagcc ggtggtctcg gcccttcatg gggccgtgcc agcccgtttc
     2761 cagaaggaca tctgtatagg gaaccagtcc aacccctgtg tgcctaacaa caaaaacccc
     2821 agggctttca atggctccag caatgggcat gtttatgaga aactttctag tattgagtct
     2881 gatgtctgta cagactacaa agaccatgac ggtgattata aagatcatga catcgactac
     2941 aaggatgacg atgacaagta actcgagata atcaacctct ggattacaaa atttgtgaaa
     3001 gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa
     3061 tgcctttgta tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat
     3121 cctggttagt tcttgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag
     3181 gggctcggct gttgggcact gacaattccg tggtgtttat ttgtgaaatt tgtgatgcta
     3241 ttgctttatt tgtaaccatc tagctttatt tgtgaaattt gtgatgctat tgctttattt
     3301 gtaaccatta taagctgcaa taaacaagtt aacaacaaca attgcattca ttttatgttt
     3361 caggttcagg gggagatgtg ggaggttttt taaagatctg gagaccggtg tggcacgtgc
     3421 ggaccgagcg gccgcaggaa cccctagtga tggagttggc cactccctct ctgcgcgctc
     3481 gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg
     3541 cctcagtgag cgagcgagcg cgcagctgcc tgcaggggcg cctgatgcgg tattttctcc
     3601 ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag caaccatagt acgcgccctg
     3661 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     3721 cagcgcctta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     3781 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
     3841 gcacctcgac cccaaaaaac ttgatttggg tgatggttca cgtagtgggc catcgccctg
     3901 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     3961 ccaaactgga acaacactca actctatctc gggctattct tttgatttat aagggatttt
     4021 gccgatttcg gtctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt
     4081 taacaaaata ttaacgttta caattttatg gtgcactctc agtacaatct gctctgatgc
     4141 cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg
     4201 tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca
     4261 gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt
     4321 tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg
     4381 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     4441 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat
     4501 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
     4561 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg
     4621 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
     4681 ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
     4741 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
     4801 ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
     4861 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc
     4921 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
     4981 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
     5041 aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca
     5101 acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
     5161 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
     5221 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
     5281 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
     5341 taagcattgg taactgtcag accaagttta ctcatatata ctttagattg atttaaaact
     5401 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
     5461 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     5521 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     5581 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     5641 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
     5701 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     5761 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     5821 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     5881 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     5941 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     6001 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     6061 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     6121 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgt
//
