Generation and Inheritance of Targeted Mutations in Potato (Solanum tuberosum L.) Using the CRISPR/Cas System

• DOI: 10.1371/journal.pone.0144591
• Publication Date: 2015-12-14T19:46:29Z
• Authors: Nathaniel M. Butler; Paul A. Atkins; Daniel F. Voytas; David S. Douches
• Keywords: Zinc finger nuclease

Genome editing using sequence-specific nucleases (SSNs) offers an alternative approach to conventional genetic engineering and opportunity extend the benefits of in agriculture. Currently available SSN platforms, such as zinc finger (ZFNs), transcription activator-like effector (TALENs), CRISPR/Cas (clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated systems (Cas)) have been used a range plant species for targeted mutagenesis via non-homologous end joining (NHEJ) are just beginning be explored crops potato (Solanum tuberosum Group Tuberosum L.). In this study, reagents expressing one two single-guide RNA (sgRNA) targeting ACETOLACTATE SYNTHASE1 (StALS1) gene were tested inducing mutations callus stable events diploid tetraploid Agrobacterium-mediated transformation with either T-DNA or modified geminivirus T-DNA. The percentage primary ranged from 3–60% per 0–29% above expected threshold based on number ALS alleles. Primary mutation frequencies cloning inheritance studies clonal propagation crosses selfing. Four nine had more than type, eight contained that maintained across generations. Somatic most evident background three four having types at single locus. Conversely, background, five candidates carried only type. Single inherited through germline both transmission percentages ranging 87–100%. This demonstration extends provides framework future studies.