LOCUS       sequence-479103-        7724 bp    DNA     circular SYN 21-JUL-2025
DEFINITION  OMEGAOff + omegaRNA scaffold AAV expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7724)
  AUTHORS   Kannan S, Altae-Tran H, Zhu S, Xu P, Strebinger D, Oshiro R, Faure 
            G, Moeller L, Pham J, Mears KS, Ni HM, Macrae RK, Zhang F
  TITLE     Evolution-guided protein design of IscB for persistent epigenome 
            editing in vivo.
  JOURNAL   Nat Biotechnol. 2025 May 7. doi: 10.1038/s41587-025-02655-3.
  PUBMED    40335752
REFERENCE   2  (bases 1 to 7724)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 21, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Biotechnol. 2025 May 7. doi: 10.1038/s41587-025-02655-3."
COMMENT     Sequence Label: pHS2056
FEATURES             Location/Qualifiers
     source          1..7724
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(71..90)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     281..302
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(456..475)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     575..597
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(635..653)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        721..825
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(826..894,895..1686)
                     /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(1044..1063)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1857..2445
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2346..2365
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   2507..2636
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        2692..2903
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      2917..2926
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2932..2952
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             6712..6732
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             6784..6969
                     /codon_start=1
                     /product="Kruppel-associated box (KRAB) transcriptional 
                     repression domain from the human zinc finger protein ZNF10 
                     (Margolin et al., 1994)"
                     /label=KRAB
                     /translation="RTLVTFKDVFVDFTREEWKLLDTAQQIVYRNVMLENYKNLVSLGY
                     QLTKPDVILRLEKGEEP"
     CDS             6976..7002
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     polyA_signal    7012..7060
                     /note="synthetic polyadenylation signal"
     promoter        complement(7236..7484)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(7294..7313)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(7464..7484)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     repeat_region   7493..7633
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   7493..7622
                     /label=AAV2 ITR
     rep_origin      join(7708..7724,1..439)
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
ORIGIN
        1 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
       61 ttagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
      121 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
      181 gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
      241 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
      301 ggaacaacac tcaactctat ctcgggctat tcttttgatt tataagggat tttgccgatt
      361 tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
      421 atattaacgt ttacaatttt atggtgcact ctcagtacaa tctgctctga tgccgcatag
      481 ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
      541 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
      601 tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag
      661 gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg
      721 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
      781 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
      841 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
      901 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
      961 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
     1021 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg
     1081 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
     1141 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     1201 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     1261 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     1321 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
     1381 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     1441 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     1501 ggctggttta ttgctgataa atctggagcc ggtgagcgtg gaagccgcgg tatcattgca
     1561 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     1621 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     1681 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     1741 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     1801 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     1861 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     1921 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     1981 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     2041 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     2101 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     2161 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     2221 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     2281 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     2341 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     2401 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     2461 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgtcctg caggcagctg
     2521 cgcgctcgct cgctcactga ggccgcccgg gcgtcgggcg acctttggtc gcccggcctc
     2581 agtgagcgag cgagcgcgca gagagggagt ggccaactcc atcactaggg gttcctgcgg
     2641 cctctagata ggtcttgaaa ggagtgggaa ttggctccgg tgcccgtcag tgggcagagc
     2701 gcacatcgcc cacagtcccc gagaagttgg ggggaggggt cggcaattga tccggtgcct
     2761 agagaaggtg gcgcggggta aactgggaaa gtgatgtcgt gtactggctc cgcctttttc
     2821 ccgagggtgg gggagaaccg tatataagtg cagtagtcgc cgtgaacgtt ctttttcgca
     2881 acgggtttgc cgccagaaca caggaaactt aagcttgcca ccatgggatc tccaaaaaag
     2941 aaacggaagg tgggatccaa ccatgaccag gaatttgacc ccccaaaggt ttacccacct
     3001 gtgccagctg agaagaggaa gcccatccgc gtgctgtctc tctttgatgg gattgctaca
     3061 gggctcctgg tgctgaagga cctgggcatc caagtggacc gctacattgc ctccgaggtg
     3121 tgtgaggact ccatcacggt gggcatggtg cggcaccagg gaaagatcat gtacgtcggg
     3181 gacgtccgca gcgtcacaca gaagcatatc caggagtggg gcccattcga cctggtgatt
     3241 ggaggcagtc cctgcaatga cctctccatt gtcaaccctg cccgcaaggg actttatgag
     3301 ggtactggcc gcctcttctt tgagttctac cgcctcctgc atgatgcgcg gcccaaggag
     3361 ggagatgatc gccccttctt ctggctcttt gagaatgtgg tggccatggg cgttagtgac
     3421 aagagggaca tctcgcgatt tcttgagtct aaccccgtga tgattgacgc caaagaagtg
     3481 tctgctgcac acagggcccg ttacttctgg ggtaaccttc ctggcatgaa caggcctttg
     3541 gcatccactg tgaatgataa gctggagctg caagagtgtc tggagcacgg cagaatagcc
     3601 aagttcagca aagtgaggac cattaccacc aggtcaaact ctataaagca gggcaaagac
     3661 cagcatttcc ccgtgttcat gaacgagaag gaggacatcc tgtggtgcac tgaaatggaa
     3721 agggtgtttg gcttccccgt ccactacaca gacgtctcca acatgagccg cttggcgagg
     3781 cagagactgc tgggccgatc gtggagcgtg ccggtcatcc gccacctctt cgctccgctg
     3841 aaggaatatt ttgcttgtgt gtctagcggc aatagtaacg ctaacagccg cgggccgagc
     3901 ttcagcagcg gcctggtgcc gttaagcttg cgcggcagcc atatgggccc tatggagata
     3961 tacaagacag tgtctgcatg gaagagacag ccagtgcggg tactgagcct cttcagaaac
     4021 atcgacaagg tactaaagag tttgggcttc ttggaaagcg gttctggttc tgggggagga
     4081 acgctgaagt acgtggaaga tgtcacaaat gtcgtgagga gagacgtgga gaaatggggc
     4141 ccctttgacc tggtgtacgg ctcgacgcag cccctaggca gctcttgtga tcgctgtccc
     4201 ggctggtaca tgttccagtt ccaccggatc ctgcagtatg cgctgcctcg ccaggagagt
     4261 cagcggccct tcttctggat attcatggac aatctgctgc tgactgagga tgaccaagag
     4321 acaactaccc gcttccttca gacagaggct gtgaccctcc aggatgtccg tggcagagac
     4381 taccagaatg ctatgcgggt gtggagcaac attccagggc tgaagagcaa gcatgcgccc
     4441 ctgaccccaa aggaagaaga gtatctgcaa gcccaagtca gaagcaggag caagctggac
     4501 gccccgaaag ttgacctcct ggtgaagaac tgccttctcc cgctgagaga gtacttcaag
     4561 tatttttctc aaaactcact tcctcttgga gggccgagct ctggcgcacc cccaccaagt
     4621 ggagggtctc ctgccgggtc cccaacatct actgaagaag gcaccagcga atccgcaacg
     4681 cccgagtcag gccctggtac ctccacagaa ccatctgaag gtagtgcgcc tggttcccca
     4741 gctggaagcc ctacttccac cgaagaaggc acgtcaaccg aaccaagtga aggatctgcc
     4801 cctgggacca gcactgaacc atctgagatg agatacgtgt acgtgctgga tgtggacggc
     4861 aagcctctga tgcctacatg caggttcggc aaggtgcgga gaatgctgaa atctggccaa
     4921 gctaaggccg tggacaccct gcctttcacc atccagctga cctacaagcc cagaaccaga
     4981 atcctgcaac ctgtgaccct gggccaggcc cccggcagaa ccaatatcgg aatggccgcc
     5041 gtgcggttcg acggcaagga actgggaaga ttccactgca tcacaagaaa caaggaaatc
     5101 cctaagctga tggccgacag aatggcagcc agaaaggcct cccggagagg cgagagactg
     5161 gccagaaagc gcctggcccg gaaactgcac accaccgcca agcacctgaa tggcagaatc
     5221 ctgccaggat gtagcgagcc catcgccgtg aaggacatca tcaacacaga gagcagattc
     5281 aacaacagaa tcctgaccaa atgcaaagtg tgcggcaaga acacccctct gagaagaaac
     5341 gtgcgggaac tcctgctgga aaacatcgtg aagaacatca gactgcacgg ccctggcgga
     5401 gagatcgtgg gcctgcaccc cgacctgaag gaaacactca agagaaccat cctggaaggc
     5461 cagcagggca atatcaacaa gctgttcagg aaacttagaa aggtgtacaa gatcaccctg
     5521 aaccagaaag actggcctgg caaaaacctg accgatatcg ctaagaataa gctgccaggc
     5581 cggctgccct tctgcaagga gcacttcgcc gagaacgaga agttcaccac catcgagaag
     5641 agcaccttcc ggctgacccc taccgccacc cagctgctgc ggacccacat caacctgttt
     5701 agaaagctga gcggcatcct gcccgtgacc gacgtggccg tcgagctgaa caagttcgcc
     5761 tttatgcagc tggacaaccc tgagatgaag aagcgggaaa tcgacttctg ccacggccca
     5821 ctgtgcggca ccggcggcct ggaagccgct gttaaggagc agcaggatgg aaaatgcctg
     5881 ctgtgtggaa aagaatctat cggccactac cacgccatcg tgcctaggtc ccgtagaggc
     5941 agcaacacca tcgctaatat cgccggcctt tgtcctaagt gccacgagct cgtgcacaag
     6001 gacgccgata cagccgagtc tctgacagag atgaaaaccg gcctgatgaa gaagtatgga
     6061 ggcaccagcg tgctgaacca gatcatcccc aagctggtgg aaaccctggc cgacctgttt
     6121 cctggccact tccacgtgac aaacggctgg aacaccaagg agttcagaga gaaacatcat
     6181 ctggaaaagg atcacgacgt ggatgcttac tgcatcgcct gcagccacct gaagcctgag
     6241 gaaaccctgg tggaaactga gcctttcgag atcctgcagt tcagaaagca caacagagcc
     6301 atcattcacc accagaccga gcggacgtac aagctggacg gcgtgacagt ggctaaaaat
     6361 agaaagaaac ggatggaaca gaaaaccgac agcctggaag attggtacgt ggacatggcc
     6421 aaggaacacg gcaaaacaca ggccgatgcc atgcggagca gactgaccgt gatcaagagc
     6481 acacggtatt acaacacccc tggcagaatg atgccaggca cagttttcct gtacgagggc
     6541 aagagatacg tgatgaccgg ccagattaca aacggcaagt actacagagc ctacggccag
     6601 gagaagcgga acttccccgc tgtcaaggtg aggattctga caaagaacac cggactagtc
     6661 tttgtggcct ctggcggctc aaaaagaacc gccgacggca gcgaattcga gcccaagaag
     6721 aagaggaaag tcggatccgg cggtggcgga gggatggatg ctaagtcact aactgcctgg
     6781 tcccggacac tggtgacctt caaggatgta tttgtggact tcaccaggga ggagtggaag
     6841 ctgctggaca ctgctcagca gatcgtgtac agaaatgtga tgctggagaa ctataagaac
     6901 ctggtttcct tgggttatca gcttactaag ccagatgtga tcctccggtt ggagaaggga
     6961 gaagagcccg gatcctaccc atacgatgtt ccagattacg cttaagaatt caataaaata
     7021 tctttatttt cattacatct gtgtgttggt tttttgtgtg aaaaaagttg taagccggta
     7081 tgagcggaaa accgcacata ccgttaaagt ttccatagca tgggcatgtc tagccggagc
     7141 gcggagcttg gaatcacccc gcggcgaatg acatgtcact gctctgctca gtaacccttt
     7201 cgggttattg acttgtcttc gtcccaggaa gacatggtgt ttcgtccttt ccacaagata
     7261 tataaagcca agaaatcgaa atactttcaa gttacggtaa gcatatgata gtccatttta
     7321 aaacataatt ttaaaactgc aaactaccca agaaattatt actttctacg tcacgtattt
     7381 tgtactaata tctttgtgtt tacagtcaaa ttaattccaa ttatctctct aacagccttg
     7441 tatcgtatat gcaaatatga aggaatcatg ggaaataggc cctcgcggcc gcaggaaccc
     7501 ctagtgatgg agttggccac tccctctctg cgcgctcgct cgctcactga ggccgggcga
     7561 ccaaaggtcg cccgacgccc gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc
     7621 agctgcctgc aggggcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca
     7681 caccgcatac gtcaaagcaa ccatagtacg cgccctgtag cggc
//
