LOCUS       sequence-489084-        7104 bp    DNA     circular SYN 09-OCT-2025
DEFINITION  5' AAVLINK plasmid for RALGAPB expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7104)
  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  JOURNAL   Unpublished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 7104)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 9, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pAAVLINK-EF1alpha-HA-5'-RALGAPB
FEATURES             Location/Qualifiers
     source          1..7104
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             36..44
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     CDS             complement(686..712)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     LTR             749..1017
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     promoter        complement(1030..1241)
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     repeat_region   1286..1415
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(1477..2065)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1557..1576)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2236..3027,3028..3096))
                     /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     2859..2878
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3097..3201)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3269..3287
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3325..3347)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3447..3466
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(3483..3938)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3620..3641)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3832..3851
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   4013..4153
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4024..4153
                     /label=AAV2 ITR
ORIGIN
        1 tccagttagc caccatctcc ttggctgttt tccaataagg aggtgttggg aagcaccgag
       61 tacaagctag taaccacacc tcaaagagaa caccaatcaa cttctctgct agattctcag
      121 caatgccacc ttgaacagtt ggtggggcca gaagtatgtc gttaatctgc agaagaaaca
      181 acagtaagac ttcccaagtt tctcgggcca tgagagatga ctcacgggcc agtttctgaa
      241 tggctctcag gacctgtaag catagtcgaa tctgactgga accctgttcc tgtcttggta
      301 caaaaagatt ctgtaggtgt tttagtatag tttgaacata ttgattaggc tctttaataa
      361 ctggcaatgg aatagaatct tttggcaaca ctaaagccat aatccagtct gtatatacat
      421 caacgcaata ttttacagtc tctccatcca atggaagggt cagtccatag caaattactt
      481 ccatggtcca ttttacttct ttgtcagttt ttaacaaatt ctcactacca gccactgaag
      541 gggtacctaa cacctgccca agaggacgga ctacagcatt tgccacctct cgtccaacgc
      601 tctctggata gctgtgcagc acactggtat ggccttgatc attctgaatc accaaatgca
      661 gtgacctcca ctcagagtaa gatccagcat agtcaggtac gtcataaggg tacatggtgg
      721 ccgtacggta ggcgccggtc acagcttgga tctgtaacgg cgcagaacag aaaacgaaac
      781 aaagacgtag agttgagcaa gcagggtcag gcaaagcgtg gagagccggc tgagtctagg
      841 taggctccaa gggagcgccg gacaaaggcc cggtctcgac ctgagcttta aacttaccta
      901 gacggcggac gcagttcagg aggcaccaca ggcgggaggc ggcagaacgc gactcaaccg
      961 gcgtggatgg cggcctcagg tagggcggcg ggcgcgtgaa ggagagatgc gagcccctcg
     1021 aagcttcagc tgtgttctgg cggcaaaccc gttgcgaaaa agaacgttca cggcgactac
     1081 tgcacttata tacggttctc ccccaccctc gggaaaaagg cggagccagt acacgacatc
     1141 actttcccag tttaccccgc gccaccttct ctaggcaccc gttcaattgc cgacccctcc
     1201 ccccaacttc tcggggactg tgggcgatgt gcgctctgcc cactgacggg caccggagcg
     1261 atcgcagatc cacgcgtgcg gccgcaggaa cccctagtga tggagttggc cactccctct
     1321 ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg cccgggcggc
     1381 ctcagtgagc gagcgagcgc gcagctgcct gcaggacatg tgagcaaaag gccagcaaaa
     1441 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     1501 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     1561 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     1621 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     1681 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     1741 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     1801 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     1861 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     1921 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     1981 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     2041 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     2101 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     2161 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     2221 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     2281 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     2341 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     2401 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     2461 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     2521 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     2581 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     2641 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     2701 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     2761 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     2821 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     2881 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     2941 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     3001 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     3061 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     3121 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     3181 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac
     3241 cattattatc atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtctcgc
     3301 gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc
     3361 ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg
     3421 cgggtgtcgg ggctggctta actatgcggc atcagagcag attgtactga gagtgcacca
     3481 taaaattgta aacgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
     3541 attttttaac caatagaccg aaatcggcaa aatcccttat aaatcaaaag aatagcccga
     3601 gatagagttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
     3661 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
     3721 caaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
     3781 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
     3841 agcgaaagga gcgggcgcta aggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
     3901 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtac tatggttgct ttgacgtatg
     3961 cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc gcatcaggcg cccctgcagg
     4021 cagctgcgcg ctcgctcgct cactgaggcc gcccgggcaa agcccgggcg tcgggcgacc
     4081 tttggtcgcc cggcctcagt gagcgagcga gcgcgcagag agggagtggc caactccatc
     4141 actaggggtt cctgcggccg ctcggtccgc acgtgccaca ccggtctcca taacttaata
     4201 acttcgtata aagtatccta tacgaataat agcttattgg tctccttaaa cctgtcttgt
     4261 aaccttgata cttacctggc ttcatataga aaataaaaac tgtgtcaaaa ggacgacatg
     4321 gcaagagatc cagataccca atgtcatcaa aaaatccagg tatcgtggaa tcaagtgcaa
     4381 taaggtgagg aggtagacga ctatttgcag gttccttcag tgcttctaag gacaaaaatc
     4441 caaagtgtga gagaaaaagg cgggctgttt ggaattcctg ggcaggaggc gggggcttgc
     4501 aatccgtaac tgggtcagga aaacttctct tctgcaattc ctcctcactc tgttgctcaa
     4561 ggtgtatttc ataagcaatc tgctgggcca tgccactcct taatttttcg tgtctctctt
     4621 ctaattcttc agtgactatt tcatgcaaat ctggaatgct gagatctgct ttcacaaaag
     4681 gaatcttgtc cacctcttca ggaaatggcc gatgtttcac agaatattta aatccaacgt
     4741 catttttagg aactgggcga ggttcaggta caaaaagctt ctgatttgct tttgctcctc
     4801 tgggtaaaag acaaagctgt tgtgcccaag caagtcttcc agacattccc cggaccagca
     4861 cagtgacaga ggggaaaaaa tcattctgct catttccaag aggttgttcc agcattgcca
     4921 ggatgacact gttatccaag acaaagtacc ggaaactatg cttgtttatg gttggcagcc
     4981 tggagtattt aatcaaagtg gtctcattca caagactaca aggagaggca ggaccactag
     5041 gtgaaggaaa tgcgccgagc aactgcataa tgcatgttag ggtggcttca gcagcatcct
     5101 ttaccctcat agatgcaggg tttggctcct tatctccttt gtacttgacc tcttgctcat
     5161 tgttcttgga cttacttcct gagataccca gttccacaat ctccagtact tccttaaggc
     5221 agtccttttc atcaagcata tcagggtgct ctgtcagcca gacacagaga cactgaaaag
     5281 ctgccactat catggagtgc agatccctgg agtgtaaagg agctggccga ctacactgat
     5341 aaacaatgta ggtgcacaca gaactgatgg ctcgcttccg gtctcctgag tcaaccatca
     5401 cttttacctt tgcaaggcca gagaggagct ctagagctgc cagtgatatg ctcatgtctt
     5461 ggcgccactg ggagttgagt ctttgggtga cgagatgaat gctgcgaatc aggagcccag
     5521 cagctgaatc tgtattaaga gcataatctc ttaagagagc ttgagcaggt ctctcactat
     5581 cgggagtcgt gggctccgtg cttccaccac ttgctgaact aataccacta ttggtgcgac
     5641 tatgactctt caggttattt tctccaccac ccatctccat ctggcaacca atggcttcca
     5701 aaagtgctga atcttgaaca atatttaaca ttgcccctaa tatcatttgg gtgttgttgg
     5761 ggtccgtttc agtttgcaag gcacctatta atatattcac aagtctcaac ttcagggaca
     5821 gaaaagttat tggtttatca taagaagagc tgtcatcgtt actaaacttt ccttccagga
     5881 ccacctcaga tttgactgtg ccaaaatgat gagggagggg caacaaagaa agcaggatat
     5941 taatggagga tcttcgcaat tctgttggat ttacatagct tttgaatttt gagagttctc
     6001 tgtcaggcaa aatggtttca agagctgaaa taaagtaagg aaccacaaca tcaatccctt
     6061 tcaagtcaca gcagaacaaa ggaggagagt ttagaataac gctggccaag acaggatggc
     6121 acacataatc atttatctgc aaaccttgaa ttaaaagcat gtaaaatctg gataaataag
     6181 ctggcagaat ctcttctcca gtcttcttgc tacaaaaaat cctacacagt gtcccacaag
     6241 cctcagctct tcctgcttca tagttatcag gaaattcact tgcatcaaac ataggattgg
     6301 aaaccatggt gtctgtggac atttgtgacc ctttccgtcg aaactccatg ctagcttgtg
     6361 ttgtaatggc agtcatgctg ctgtctctgt ttatcccatt atgaagttta cagtgaacaa
     6421 atgctgcatc aaataaccat gatccaaaga gattcaagat gctgttaacc tttggtctcc
     6481 gaggggcagg cagtggccgg ggttctgaac tagtctgatt tggacttgac agaggagagg
     6541 tggaggacgt ctggtgctga actttagagg catgagcagt actaactgtg cttgtcttca
     6601 tggtggcctt attcacagta acagcccggt gcctccggtt atgtgggggg gtggtactga
     6661 cagcagcact ttgaggcata cttaatctat tcacgggtgt tgggggagca ctgtctgatc
     6721 ggggtctaga aatacctaag aatgcatcca ccagacagct gattccacgc atggcacgaa
     6781 aaaatatttg aggcagatgt ttaaggcagg gatactgatt caattcttgc ggcattccgc
     6841 tcacattcaa gaactgttcc tgaaatttgg gagtagagct tataatagct gggttactca
     6901 aatccacagg attacttaac atgtgtaaaa agcgaaacca tgtctgtgca acacactcat
     6961 tatccatttc tggagggatc agactggcat cttcatcggg aactttaaat gcaggaaatg
     7021 aaggaccata tgtaaagcgt agcaatctgg aagtgagtgc acaaatgacc ttgctccact
     7081 gctccaccac tgctgggtga tgcc
//
