Novel guide rna/cas endonuclease systems

Abstract

Compositions and methods are provided for novel guide RNA / Cas endonuclease systems. Type II Cas9 endonuclease systems originating from Brevibacillus laterosporus, Lactobacillus reuteri Mlc3, Lactobacillus rossiae DSM 15814, Pediococcus pentosaceus SL4, Lactobacillus nodensis JCM 14932, Sulfurospirillum sp. SCADC, Bifidobacterium thermophilum DSM 20210, Loktanella vestfoldensis, Sphingomonas sanxanigenens NX02, Epilithonimonas tenax DSM 16811, Sporocytophaga myxococcoides are described herein. The present disclosure also describes methods for genome modification of a target sequence in the genome of a cell, for gene editing, and for inserting a polynucleotide of interest into the genome of a cell.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS[0001]This application is a continuation of U.S. patent application Ser. No. 15 / 573,953, filed 14 Nov. 2017, which is the 371 national stage entry of International Application Number PCT / US2016 / 032073, filed on 12 May 2016, which claims the benefit of U.S. Provisional Application No. 62 / 162,377, filed May 15, 2015, U.S. Provisional Application No. 62 / 162,353, filed May 15, 2015 and U.S. Provisional Application No. 62 / 196,535, filed Jul. 24, 2015, each of which is incorporated herein in their entirety by reference.FIELD[0002]The disclosure relates to the field of plant molecular biology, in particular, to compositions for novel guide RNA / Cas endonuclease systems and compositions and methods for altering the genome of a cell.REFERENCE TO SEQUENCE LISTING SUBMITTED ELECTRONICALLY[0003]The official copy of the sequence listing is submitted electronically via EFS-Web as an ASCII formatted sequence listing with a file named BB2539USPCN_SequenceListin...

AI Technical Summary

Benefits of technology

The patent text describes methods and compositions for quickly characterizing new Cas endonuclease systems and their elements. These methods involve using guide RNAs that can recognize and bind to target sequences, and optionally nick or cleave them. The guide RNAs can be duplex molecules comprising a chimeric non-naturally occurring crRNA and a tracrRNA, or single molecules comprising a chimeric non-naturally occurring crRNA and a tracrRNA. The methods can be used with various organisms, such as Brevibacillus laterosporus, Lactobacillus reuteri, and Bifidobacterium thermophilum. The patent also provides nucleic acid constructs, plants, plant cells, explants, seeds, and grain with altered target sites or polynucleotides of interest produced by the methods described herein.

Problems solved by technology

Genome-editing techniques such as designer zinc finger nucleases (ZFNs) or transcription activator-like effector nucleases (TALENs), or homing meganucleases, are available for producing targeted genome perturbations, but these systems tends to have a low specificity and employ designed nucleases that need to be redesigned for each target site, which renders them costly and time-consuming to prepare.

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