LOCUS       sequence-491888-        6439 bp    DNA     circular SYN 04-NOV-2025
DEFINITION  3' AAVLINK plasmid for CLTCL1 expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6439)
  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  JOURNAL   Unpublished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 6439)
  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            "Unpublished"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'-CLTCL1-FLAG
FEATURES             Location/Qualifiers
     source          1..6439
                     /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             3159..3224
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(3289..3309)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   3702..3842
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3702..3831
                     /label=AAV2 ITR
     rep_origin      3917..4372
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4004..4023)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4214..4235
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4389..4408)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4508..4530
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4568..4586)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4654..4758
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4759..4827,4828..5619)
                     /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(4977..4996)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5790..6378
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6279..6298
                     /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 ggtcctgatc gagcacattg
     1501 gaaacctgga ccgggcatat gagtttgcgg agagatgcaa tgagcctgct gtgtggagtc
     1561 agctggccca agcccagctc cagaaagatt tggtgaagga agccatcaac tcctatatca
     1621 gaggggacga cccttcctct tacctggaag ttgttcagtc agccagcagg agcaacaact
     1681 gggaggatct agttaaattt ctgcagatgg ccaggaaaaa gggccgtgag tcctatatag
     1741 agactgaact tatttttgcc ttggctaaaa ccagccgtgt ttctgagcta gaagatttta
     1801 ttaatggacc caacaatgcc cacatccagc aggttggaga ccgctgttac gaggagggaa
     1861 tgtacgaggc tgccaagctg ctctatagca atgtttctaa ctttgcccgc ccggcttcca
     1921 ccttggttca cctcggtgag tatcaggcag cagtggacaa cagccgcaag gccagcagca
     1981 cccggacgtg gaaggaggtg tgctttgcct gcatggatgg acaagagttc cgcttcgcac
     2041 agctgtgtgg tcttcacatc gtcattcatg cagatgagct ggaggagctg atgtgctatt
     2101 accaggatcg tggctacttt gaggagctga tcttgctgtt ggaagcggcc ctgggcctgg
     2161 agcgggccca catgggcatg ttcactgagc tggccatcct ctactccaaa ttcaagccac
     2221 agaagatgct ggagcatctg gagcttttct ggtcccgtgt caacatccca aaggtgctga
     2281 gggctgcaga gcaggcacac ctgtgggctg agctggtgtt cctctatgac aagtacgagg
     2341 agtatgacaa tgctgtgctc accatgatga gccaccccac tgaggcctgg aaggagggtc
     2401 agttcaagga catcattacc aaggttgcca acgtcgagct ctgttacaga gccctgcagt
     2461 tctatttgga ttacaaacca ctgctcatca atgacctgct gctggtgctt tcaccccggc
     2521 tggaccacac ctggacagtc agtttctttt caaaggcagg tcagctgccc ctggtgaagc
     2581 cttacctgcg gtcagtccag agccacaaca acaagagtgt gaatgaggca ctcaaccacc
     2641 tgctgacaga ggaggaggac tatcagggct taagggcatc tatcgatgcc tatgacaact
     2701 ttgacaacat cagcctggct cagcagctgg agaagcatca gctgatggag ttcaggtgca
     2761 ttgcggccta tctgtacaag ggcaataact ggtgggccca gagcgtggag ctctgcaaga
     2821 aggatcatct ctacaaggat gccatgcagc atgctgcaga gtcgcgggat gctgagctgg
     2881 cccagaagtt gctgcagtgg ttcctggagg aaggcaagag ggagtgcttc gcagcttgtc
     2941 tcttcacctg ctatgacctg cttcgcccag acatggtgct tgagctggcc tggaggcaca
     3001 acctcgtgga cttggccatg ccctacttca tccaggtgat gagggagtac ctgagcaagg
     3061 tggacaaact ggatgccttg gagagtctgc gcaagcaaga ggagcatgtg acagagcctg
     3121 cccctctcgt gtttgatttt gatgggcatg aatgtacaga ctacaaagac catgacggtg
     3181 attataaaga tcatgacatc gactacaagg atgacgatga caagtaactc gagataatca
     3241 acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt
     3301 tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc
     3361 tttcattttc tcctccttgt ataaatcctg gttagttctt gccacggcgg aactcatcgc
     3421 cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt
     3481 gtttatttgt gaaatttgtg atgctattgc tttatttgta accatctagc tttatttgtg
     3541 aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca
     3601 acaacaattg cattcatttt atgtttcagg ttcaggggga gatgtgggag gttttttaaa
     3661 gatctggaga ccggtgtggc acgtgcggac cgagcggccg caggaacccc tagtgatgga
     3721 gttggccact ccctctctgc gcgctcgctc gctcactgag gccgggcgac caaaggtcgc
     3781 ccgacgcccg ggctttgccc gggcggcctc agtgagcgag cgagcgcgca gctgcctgca
     3841 ggggcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatacg
     3901 tcaaagcaac catagtacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
     3961 acgcgcagcg tgaccgctac acttgccagc gccttagcgc ccgctccttt cgctttcttc
     4021 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
     4081 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga tttgggtgat
     4141 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
     4201 acgttcttta atagtggact cttgttccaa actggaacaa cactcaactc tatctcgggc
     4261 tattcttttg atttataagg gattttgccg atttcggtct attggttaaa aaatgagctg
     4321 atttaacaaa aatttaacgc gaattttaac aaaatattaa cgtttacaat tttatggtgc
     4381 actctcagta caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca
     4441 cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg
     4501 accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga
     4561 cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct
     4621 tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
     4681 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
     4741 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt
     4801 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct
     4861 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc
     4921 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta
     4981 tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac
     5041 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc
     5101 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac
     5161 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg
     5221 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac
     5281 gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc
     5341 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt
     5401 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga
     5461 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc
     5521 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag
     5581 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca
     5641 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     5701 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     5761 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     5821 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     5881 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt
     5941 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     6001 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     6061 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     6121 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     6181 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     6241 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     6301 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     6361 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     6421 tggccttttg ctcacatgt
//
