LOCUS       sequence-372789-        3838 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Single guide RNA cassette under U6 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3838)
  AUTHORS   Bieluszewski T, Sura W, Dziegielewski W, Bieluszewska A, Lachance C,
            Kabza M, Szymanska-Lejman M, Abram M, Wlodzimierz P, De Winne N, De 
            Jaeger G, Sadowski J, Cote J, Ziolkowski PA
  TITLE     NuA4 and H2A.Z control environmental responses and autotrophic 
            growth in Arabidopsis.
  JOURNAL   Nat Commun. 2022 Jan 12;13(1):277. doi: 10.1038/s41467-021-27882-5.
  PUBMED    35022409
REFERENCE   2  (bases 1 to 3838)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2022-01-12"; volume: "13(1):277. doi"; pages: " 
            10.1038/s41467-021-27882-5"
COMMENT     Sequence Label: pJET1.2-U6
FEATURES             Location/Qualifiers
     source          1..3838
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(3..20)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(174..762)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(254..273)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(933..1724,1725..1793))
                     /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     1556..1575
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1794..1898)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1966..1984
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2290..2308
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     2356..2373
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2357..2373
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2380..2398
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_RNA        2930..3005
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(3112..3128)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3112..3128)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     promoter        complement(3165..3183)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3204..3220)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3204..3220)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        complement(join(3655..3661,3662..3679,3680..3685))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    3700..3721
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
       61 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
      121 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
      181 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
      241 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
      301 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
      361 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
      421 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
      481 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
      541 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
      601 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
      661 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
      721 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
      781 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
      841 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
      901 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
      961 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     1021 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     1081 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     1141 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     1201 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     1261 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     1321 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     1381 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     1441 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     1501 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     1561 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     1621 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     1681 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     1741 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     1801 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     1861 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagttcca
     1921 cctgacgtct aagaaaccat tattatcatg acattaacct ataaaaatag gcgtatcacg
     1981 aggccgcccc tgcagccgaa ttatattatt tttgccaaat aatttttaac aaaagctctg
     2041 aagtcttctt catttaaatt cttagatgat acttcatctg gaaaattgtc ccaattagta
     2101 gcatcacgct gtgagtaagt tctaaaccat ttttttattg ttgtattatc tctaatctta
     2161 ctactcgatg agttttcggt attatctcta tttttaactt ggagcaggtt ccattcattg
     2221 tttttttcat catagtgaat aaaatcaact gctttaacac ttgtgcctga acaccatatc
     2281 catccggcgt aatacgactc actataggga gagcggccgc cagatcttcc ggatggctcg
     2341 agtttttcag caagatgtaa aacgacggcc agtgaattgt aatacgactc actatagggc
     2401 gaattgggta ccgagctcgg atccactagt aacggacgcc agtgtgctgg aattgccctt
     2461 aagcttcgtt gaacaacgga aactcgactt gccttccgca caatacatca tttcttctta
     2521 gctttttttc ttcttcttcg ttcatacagt ttttttttgt ttatcagctt acattttctt
     2581 gaaccgtagc tttcgttttc ttctttttaa ctttccattc ggagtttttg tatcttgttt
     2641 catagtttgt cccaggatta gaatgattag gcatcgaacc ttcaagaatt tgattgaata
     2701 aaacatcttc attcttaaga tatgaagata atcttcaaaa ggcccctggg aatctgaaag
     2761 aagagaagca ggcccattta tatgggaaag aacaatagta tttcttatat aggcccattt
     2821 aagttgaaaa caatcttcaa aagtcccaca tcgcttagat aagaaaacga agctgagttt
     2881 atatacagct agagtcgaag tagtgattgc tcgcactagt gaaccccagg ttttagagct
     2941 agaaatagca agataaaata aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc
     3001 ggtgcttttt ttgtcccttc gaagggcaat tctgcagata tccatcacac tggcggccgc
     3061 tagaggtcga cggtatcgat aagcttgata tcgaattcct gcagcccggg ggatccacta
     3121 gttctagaac ggccgccacc gcggtggagc tccagctttt gttcccttta gtgagggtta
     3181 atttcgagct tggcgtaatc atggtcatag ctgtttcctg atctttctag aagatctcct
     3241 acaatattct cagctgccat ggaaaatcga tgttcttctt ttattctctc aagattttca
     3301 ggctgtatat taaaacttat attaagaact atgctaacca cctcatcagg aaccgttgta
     3361 ggtggcgtgg gttttcttgg caatcgactc tcatgaaaac tacgagctaa atattcaata
     3421 tgttcctctt gaccaacttt attctgcatt ttttttgaac gaggtttaga gcaagcttca
     3481 ggaaactgag acaggaattt tattaaaaat ttaaattttg aagaaagttc agggttaata
     3541 gcatccattt tttgctttgc aagttcctca gcattcttaa caaaagacgt ctcttttgac
     3601 atgtttaaag tttaaacctc ctgtgtgaaa ttattatccg ctcataattc cacacattat
     3661 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     3721 aattgcgttg cgctcactgc caattgcttt ccagtcggga aacctgtcgt gccagctgca
     3781 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgct
//
