Chemically modified guide RNAS for CRISPR/Cas-mediated gene regulation

A primary cell comprising:

• (a) a modified single guide RNA (sgRNA) comprising a first nucleotide sequence that is complementary to a target nucleic acid within the primary cell and a second nucleotide sequence that interacts with a CRISPR-associated protein (Cas) polypeptide, wherein:
• (i) about 10% to about 30% of nucleotides in the first nucleotide sequence and/or about 1% to about 10% of nucleotides in the second nucleotide sequence are modified nucleotides;
• (ii) the modified sgRNA comprises a modified nucleotide within five nucleotides from the 5′ end of the first nucleotide sequence and/or within five nucleotides from the 3′ end of the second nucleotide sequence;
• and
• (iii) the modified nucleotides are selected from the group consisting of a 2′-O-methyl 3′-phosphorothioate (MS) nucleotide, a 2′-O-methyl 3′-thioPACE (MSP) nucleotide, and a combination thereof;
• and
• (b) a Cas polypeptide, an mRNA encoding the Cas polypeptide, or a recombinant expression vector comprising a nucleotide sequence encoding the Cas polypeptide,
• wherein the modified sgRNA guides the Cas polypeptide to the target nucleic acid, and wherein the modified sgRNA induces a gene regulation of the target nucleic acid with an enhanced activity relative to a corresponding unmodified sgRNA.