LOCUS       sequence-478218-        9744 bp    DNA     circular SYN 11-JUL-2025
DEFINITION  Express AAV2-KO (R585A,R588A) capsid with ncAA mutant at site 588 of
            VP1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9744)
  AUTHORS   Pham Q, Glicksman J, Shahraeini S, Han B, Jewel D, Loynd C, Singha 
            Roy SJ, Chatterjee A
  TITLE     A facile chemical strategy to synthesize precise AAV-protein 
            conjugates for targeted gene delivery
  JOURNAL   bioRxiv 2024.07.20.604406
REFERENCE   2  (bases 1 to 9744)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 11, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            2024.07.20.604406"
COMMENT     Sequence Label: pIDTsmart-RC2-KO-delVP1-VP1-588TAG
FEATURES             Location/Qualifiers
     source          1..9744
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     6..22
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    235..282
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             4231..4242
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     primer_bind     complement(4717..4733)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4726..4748)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     enhancer        4917..5296
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5297..5500
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5451..5471
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     5545..5564
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5545..5563
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             7632..7643
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     primer_bind     complement(8045..8061)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8045..8061)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     terminator      8109..8140
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     promoter        8141..8243
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             8244..9059
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCLKPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNGDRVFRLAQAQSRMNNGLVGA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(8317..8336)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      9083..9110
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     rep_origin      complement(9122..9709)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(9202..9221)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cccgtgtaaa acgacggcca gtttatctag tcagcttgat tctagctgat cgtggaccgg
       61 aaggtgagcc agtgagttga ttgcagtcca gttacgctgg agtctgaggc tcgtcctgaa
      121 tgatatgcga ccgccggagg gttgcgtttg agacgggcga cagatccagt cgcgctgctc
      181 tcgtcgatcc gctagggcgg ccgctctaga actagtggat cccccggaag atcagaagtt
      241 cctattccga agttcctatt ctctagaaag tataggaact tctgatctgc gcagccgcca
      301 tgccggggtt ttacgagatt gtgattaagg tccccagcga ccttgacgag catctgcccg
      361 gcatttctga cagctttgtg aactgggtgg ccgagaagga atgggagttg ccgccagatt
      421 ctgacatgga tctgaatctg attgagcagg cacccctgac cgtggccgag aagctgcagc
      481 gcgactttct gacggaatgg cgccgtgtga gtaaggcccc ggaggccctt ttctttgtgc
      541 aatttgagaa gggagagagc tacttccaca tgcacgtgct cgtggaaacc accggggtga
      601 aatccatggt tttgggacgt ttcctgagtc agattcgcga aaaactgatt cagagaattt
      661 accgcgggat cgagccgact ttgccaaact ggttcgcggt cacaaagacc agaaatggcg
      721 ccggaggcgg gaacaaggtg gtggatgagt gctacatccc caattacttg ctccccaaaa
      781 cccagcctga gctccagtgg gcgtggacta atatggaaca gtatttaagc gcctgtttga
      841 atctcacgga gcgtaaacgg ttggtggcgc agcatctgac gcacgtgtcg cagacgcagg
      901 agcagaacaa agagaatcag aatcccaatt ctgatgcgcc ggtgatcaga tcaaaaactt
      961 cagccaggta catggagctg gtcgggtggc tcgtggacaa ggggattacc tcggagaagc
     1021 agtggatcca ggaggaccag gcctcataca tctccttcaa tgcggcctcc aactcgcggt
     1081 cccaaatcaa ggctgccttg gacaatgcgg gaaagattat gagcctgact aaaaccgccc
     1141 ccgactacct ggtgggccag cagcccgtgg aggacatttc cagcaatcgg atttataaaa
     1201 ttttggaact aaacgggtac gatccccaat atgcggcttc cgtctttctg ggatgggcca
     1261 cgaaaaagtt cggcaagagg aacaccatct ggctgtttgg gcctgcaact accgggaaga
     1321 ccaacatcgc ggaggccata gcccacactg tgcccttcta cgggtgcgta aactggacca
     1381 atgagaactt tcccttcaac gactgtgtcg acaagatggt gatctggtgg gaggagggga
     1441 agatgaccgc caaggtcgtg gagtcggcca aagccattct cggaggaagc aaggtgcgcg
     1501 tggaccagaa atgcaagtcc tcggcccaga tagacccgac tcccgtgatc gtcacctcca
     1561 acaccaacat gtgcgccgtg attgacggga actcaacgac cttcgaacac cagcagccgt
     1621 tgcaagaccg gatgttcaaa tttgaactca cccgccgtct ggatcatgac tttgggaagg
     1681 tcaccaagca ggaagtcaaa gactttttcc ggtgggcaaa ggatcacgtg gttgaggtgg
     1741 agcatgaatt ctacgtcaaa aagggtggag ccaagaaaag acccgccccc agtgacgcag
     1801 atataagtga gcccaaacgg gtgcgcgagt cagttgcgca gccatcgacg tcagacgcgg
     1861 aagcttcgat caactacgca gacaggtacc aaaacaaatg ttctcgtcac gtgggcatga
     1921 atctgatgct gtttccctgc agacaatgcg agagaatgaa tcagaattca aatatctgct
     1981 tcactcacgg acagaaagac tgtttagagt gctttcccgt gtcagaatct caacccgttt
     2041 ctgtcgtcaa aaaggcgtat cagaaactgt gctacattca tcatatcatg ggaaaggtgc
     2101 cagacgcttg cactgcctgc gatctggtca atgtggattt ggatgactgc atctttgaac
     2161 aataaatgat ttaaatcagg tctcgctgcc gatggttatc ttccagattg gctcgaggac
     2221 actctctctg aaggaataag acagtggtgg aagctcaaac ctggcccacc accaccaaag
     2281 cccgcagagc ggcataagga cgacagcagg ggtcttgtgc ttcctgggta caagtacctc
     2341 ggacccttca acggactcga caagggagag ccggtcaacg aggcagacgc cgcggccctc
     2401 gagcacgaca aagcctacga ccggcagctc gacagcggag acaacccgta cctcaagtac
     2461 aaccacgccg acgcggagtt tcaggagcgc cttaaagaag atacgtcttt tgggggcaac
     2521 ctcggacgag cagtcttcca ggcgaaaaag agggttcttg aacctctggg cctggttgag
     2581 gaacctgtta agacggctcc gggaaaaaag aggccggtag agcactctcc tgtggagcca
     2641 gactcctcct cgggaaccgg aaaggcgggc cagcagcctg caagaaaaag attgaatttt
     2701 ggtcagactg gagacgcaga ctcagtacct gacccccagc ctctcggaca gccaccagca
     2761 gccccctctg gtctgggaac taatacgatg gctacaggca gtggcgcacc aatggcagac
     2821 aataacgagg gcgccgacgg agtgggtaat tcctcgggaa attggcattg cgattccaca
     2881 tggatgggcg acagagtcat caccaccagc acccgaacct gggccctgcc cacctacaac
     2941 aaccacctct acaaacaaat ttccagccaa tcaggagcct cgaacgacaa tcactacttt
     3001 ggctacagca ccccttgggg gtattttgac ttcaacagat tccactgcca cttttcacca
     3061 cgtgactggc aaagactcat caacaacaac tggggattcc gacccaagag actcaacttc
     3121 aagctcttta acattcaagt caaagaggtc acgcagaatg acggtacgac gacgattgcc
     3181 aataacctta ccagcacggt tcaggtgttt actgactcgg agtaccagct cccgtacgtc
     3241 ctcggctcgg cgcatcaagg atgcctcccg ccgttcccag cagacgtctt catggtgcca
     3301 cagtatggat acctcaccct gaacaacggg agtcaggcag taggacgctc ttcattttac
     3361 tgcctggagt actttccttc tcagatgctg cgtaccggaa acaactttac cttcagctac
     3421 acttttgagg acgttccttt ccacagcagc tacgctcaca gccagagtct ggaccgtctc
     3481 atgaatcctc tcatcgacca gtacctgtat tacttgagca gaacaaacac tccaagtgga
     3541 accaccacgc agtcaaggct tcagttttct caggccggag cgagtgacat tcgggaccag
     3601 tctaggaact ggcttcctgg accctgttac cgccagcagc gagtatcaaa gacatctgcg
     3661 gataacaaca acagtgaata ctcgtggact ggagctacca agtaccacct caatggcaga
     3721 gactctctgg tgaatccggg cccggccatg gcaagccaca aggacgatga agaaaagttt
     3781 tttcctcaga gcggggttct catctttggg aagcaaggct cagagaaaac aaatgtggac
     3841 attgaaaagg tcatgattac agacgaagag gaaatcagga caaccaatcc cgtggctacg
     3901 gagcagtatg gttctgtatc taccaacctc caggccggca acgcccaagc agctaccgca
     3961 gatgtcaaca cacaaggcgt tcttccaggc atggtctggc aggacagaga tgtgtacctt
     4021 caggggccca tctgggcaaa gattccacac acggacggac attttcaccc ctctcccctc
     4081 atgggtggat tcggacttaa acaccctcct ccacagattc tcatcaagaa caccccggta
     4141 cctgcgaatc cttcgaccac cttcagtgcg gcaaagtttg cttccttcat cacacagtac
     4201 tccacgggac aggtcagcgt ggagatcgag tgggagctgc agaaggaaaa cagcaaacgc
     4261 tggaatcccg aaattcagta cacttccaac tacaacaagt ctgttaatgt ggactttact
     4321 gtggacacta atggcgtgta ttcagagcct cgccccattg gcaccagata cctgactcgt
     4381 aatctgtaat tgcttgttaa tcaataaacc gtttaattcg tttcagttga actttggtct
     4441 ctgcgtattt ctttcttatc tagtttccat ggctacgtag ataagtagca tggcgggtta
     4501 atcattaact acagcccggg cgtttaaaca gcgggcggag gggtggagtc gtgacgtgaa
     4561 ttacgtcata gggttaggga ggtcctgtat tagaggtcac gtgagtgttt tgcgacattt
     4621 tgcgacacca tgtggtctcg ctgggggggg gggcccgagt gagcacgcag ggtctccatt
     4681 ttgaagcggg aggtttgaac gagcgctggc gcgctcactg gccgtcgttt tacaacgtcg
     4741 tgactgggaa aaccctggcg ttacccaact taatcgcctt gcagcacatc cccctttcgc
     4801 cagctggcgt aatagcgaag aggcccgcac cgatcgccct tcccatgcat cggccgcaaa
     4861 tacctgcagg atccgttttg cgctgcttcg cgatgtacgg gccagatata cgcgttgaca
     4921 ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc atagcccata
     4981 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
     5041 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
     5101 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
     5161 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
     5221 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
     5281 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
     5341 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
     5401 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
     5461 cggtaggcgt gtacggtggg aggtctatat aagcagagct ctctggctaa ctagagaacc
     5521 cactgcttac tggcttatcg aaattaatac gactcactat agggagaccc aagctggcta
     5581 gcatggctgc cgatggttat cttccagatt ggctcgagga cactctctct gaaggaataa
     5641 gacagtggtg gaagctcaaa cctggcccac caccaccaaa gcccgcagag cggcataagg
     5701 acgacagcag gggtcttgtg cttcctgggt acaagtacct cggacccttc aacggactcg
     5761 acaagggaga gccggtcaac gaggcagacg ccgcggccct cgagcacgac aaagcctacg
     5821 accggcagct cgacagcgga gacaacccgt acctcaagta caaccacgcc gacgcggagt
     5881 ttcaggagcg ccttaaagaa gatacgtctt ttgggggcaa cctcggacga gcagtcttcc
     5941 aggcgaaaaa gagggttctt gaacctctgg gcctggttga ggaacctgtt aagaccgctc
     6001 cgggaaaaaa gaggccggta gagcactctc ctgtggagcc agactcctcc tcgggaaccg
     6061 gaaaggcggg ccagcagcct gcaagaaaaa gattgaattt tggtcagact ggagacgcag
     6121 actcagtacc tgacccccag cctctcggac agccaccagc agccccctct ggtctcggaa
     6181 ctaataccct cgctacaggc agtggcgcac cactcgcaga caataacgag ggcgccgacg
     6241 gagtgggtaa ttcctcggga aattggcatt gcgattccac atggctcggc gacagagtca
     6301 tcaccaccag cacccgaacc tgggccctgc ccacctacaa caaccacctc tacaaacaaa
     6361 tttccagcca atcaggagcc tcgaacgaca atcactactt tggctacagc accccttggg
     6421 ggtattttga cttcaacaga ttccactgcc acttttcacc acgtgactgg caaagactca
     6481 tcaacaacaa ctggggattc cgacccaaga gactcaactt caagctcttt aacattcaag
     6541 tcaaagaggt cacgcagaat gacggtacga cgacgattgc caataacctt accagcacgg
     6601 ttcaggtgtt tactgactcg gagtaccagc tcccgtacgt cctcggctcg gcgcatcaag
     6661 gatgcctccc gccgttccca gcagacgtct tcatggtgcc acagtatgga tacctcaccc
     6721 tgaacaacgg gagtcaggca gtaggacgct cttcatttta ctgcctggag tactttcctt
     6781 ctcagatgct gcgtaccgga aacaacttta ccttcagcta cacttttgag gacgttcctt
     6841 tccacagcag ctacgctcac agccagagtc tggaccgtct catgaatcct ctcatcgacc
     6901 agtacctgta ttacttgagc agaacaaaca ctccaagtgg aaccaccacg cagtcaaggc
     6961 ttcagttttc tcaggccgga gcgagtgaca ttcgggacca gtctaggaac tggcttcctg
     7021 gaccctgtta ccgccagcag cgagtatcaa agacatctgc ggataacaac aacagtgaat
     7081 actcgtggac tggagctacc aagtaccacc tcaatggcag agactctctg gtgaatccgg
     7141 gcccggccat ggcaagccac aaggacgatg aagaaaagtt ttttcctcag agcggggttc
     7201 tcatctttgg gaagcaaggc tcagagaaaa caaatgtgga cattgaaaag gtcatgatta
     7261 cagacgaaga ggaaatcagg acaaccaatc ccgtggctac ggagcagtat ggttctgtat
     7321 ctaccaacct ccagagaggc aactagcaag cagctaccgc agatgtcaac acacaaggcg
     7381 ttcttccagg catggtctgg caggacagag atgtgtacct tcaggggccc atctgggcaa
     7441 agattccaca cacggacgga cattttcacc cctctcccct catgggtgga ttcggactta
     7501 aacaccctcc tccacagatt ctcatcaaga acaccccggt acctgcgaat ccttcgacca
     7561 ccttcagtgc ggcaaagttt gcttccttca tcacacagta ctccacggga caggtcagcg
     7621 tggagatcga gtgggagctg cagaaggaaa acagcaaacg ctggaatccc gaaattcagt
     7681 acacttccaa ctacaacaag tctgttaatg tggactttac tgtggacact aatggcgtgt
     7741 attcagagcc tcgccccatt ggcaccagat acctgactcg taatctgtaa ttgcttgtta
     7801 atcaataaac cgtttaattc gtttcagttg aactttggtc tctgcgtatt tctttcttat
     7861 ctagtttcca tggctacgta gataagtagc atggcgggtt aatcattaac tacagcccta
     7921 ggggtgcgag cggatcgagc agtgtcgatc actactggac cgcgagctgt gctgcgaccc
     7981 gtgatcttac ggcattatac gtatgatcgg tccacgatca gctagattat ctagtcagct
     8041 tgatgtcata gctgtttcct gaggctcaat actgaccatt taaatcatac ctgacctcca
     8101 tagcagaaag tcaaaagcct ccgaccggag gcttttgact tgatcggcac gtaagaggtt
     8161 ccaactttca ccataatgaa ataagatcac taccgggcgt attttttgag ttatcgagat
     8221 tttcaggagc taaggaagct aaaatgagcc atattcaacg ggaaacgtct tgcttgaagc
     8281 cgcgattaaa ttccaacatg gatgctgatt tatatgggta taaatgggct cgcgataatg
     8341 tcgggcaatc aggtgcgaca atctatcgat tgtatgggaa gcccgatgcg ccagagttgt
     8401 ttctgaaaca tggcaaaggt agcgttgcca atgatgttac agatgagatg gtcaggctaa
     8461 actggctgac ggaatttatg cctcttccga ccatcaagca ttttatccgt actcctgatg
     8521 atgcatggtt actcaccact gcgatcccag ggaaaacagc attccaggta ttagaagaat
     8581 atcctgattc aggtgaaaat attgttgatg cgctggcagt gttcctgcgc cggttgcatt
     8641 cgattcctgt ttgtaattgt ccttttaacg gcgatcgcgt atttcgtctc gctcaggcgc
     8701 aatcacgaat gaataacggt ttggttggtg cgagtgattt tgatgacgag cgtaatggct
     8761 ggcctgttga acaagtctgg aaagaaatgc ataaactctt gccattctca ccggattcag
     8821 tcgtcactca tggtgatttc tcacttgata accttatttt tgacgagggg aaattaatag
     8881 gttgtattga tgttggacga gtcggaatcg cagaccgata ccaggatctt gccatcctat
     8941 ggaactgcct cggtgagttt tctccttcat tacagaaacg gctttttcaa aaatatggta
     9001 ttgataatcc tgatatgaat aaattgcagt ttcacttgat gctcgatgag tttttctaat
     9061 gaggacctaa atgtaatcac ctggctcacc ttcgggtggg cctttctgcg ttgctggcgt
     9121 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgatgctca agtcagaggt
     9181 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     9241 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     9301 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     9361 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     9421 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     9481 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     9541 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     9601 cctcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     9661 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     9721 gattttctac cgaagaaagg ccca
//
