Switchable gRNAs comprising aptamers

A switchable single-guide RNA (sgRNA), comprising

• (1) an sgRNA backbone sequence, wherein the sgRNA backbone sequence comprises a nucleotide sequence that is at least 90% identical to the nucleotide sequence 5′- GUUUUAGAGC UAUGCUGAAA AGCAUAGCAA GUUAAAAUAA GGCUAGUCCG UUAUC -3′(nucleotides 79-133 of SEQ ID NO:
• 6) and/or at least 90% identical to the nucleotide sequence 5′- GUUUUAGAGC UAUGCUGAAA AGCAUAGCAA GUUAAAAU -3′(nucleotides 22-59 of SEQ ID NO:
• 9);
• (2) an RNA aptamer comprising a riboswitch, wherein the aptamer comprises 20-100 nucleotides, and wherein the riboswitch is selected from a theophylline riboswitch, a thiamine pyrophosphate (TPP) riboswitch, an adenosine cobalamin (AdoCbl) riboswitch, an S-adenosyl methionine (SAM) riboswitch, an SAH riboswitch, a flavin mononucleotide (FMN) riboswitch, a tetrahydrofolate riboswitch, a lysine riboswitch, a glycine riboswitch, a purine riboswitch, a GlmS riboswitch, or a pre-queosinel (PreQ1) riboswitch;
• and
• (3) a guide sequence that is complementary to a DNA target site,
• wherein the DNA target site comprises the structure 5′-[Nz]-[protospacer adjacent motif (PAM)]-3′, wherein
• each N is, independently, any nucleotide, and
• Z is 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, or 50;
• wherein the guide sequence of (3) is flanked by the riboswitch of (2) on one side and by the sgRNA backbone sequence of (1) on the other side;
• and
• wherein the riboswitch of (2) comprises a sequence of at least 4 contiguous nucleotides that is complementary to the guide sequence.