Selection of sugarcane introns with low propensity to form stable secondary structures

• Grant number: 23/02452-8
• Support Opportunities: Scholarships abroad - Research Internship - Scientific Initiation
• Start date: June 01, 2023
• End date: July 31, 2023
• Field of knowledge: Biological Sciences - Biology
• Principal Investigator: Diego Mauricio Riano Pachon
• Grantee: Beatriz Rodrigues Estevam
• Supervisor: Dirk Walther
• Host Institution: Centro de Energia Nuclear na Agricultura (CENA). Universidade de São Paulo (USP). Piracicaba , SP, Brazil
• Institution abroad: Max Planck Society, Potsdam, Germany
• Associated to the scholarship: 22/10264-4 - In silico selection of sugarcane introns to aid in genome editing, BP.IC

Abstract

Plant breeding to produce desired characteristics is a common practice and it is in constant expansion. Recently, CRISPR/Cas technology has been used to introduce highly specific mutations in plant genomes and has been increasingly implemented in breeding-programs in Brazil. Sugarcane (Saccharum spp.) is an important economic crop for energy and food production. However, due to their complex genome, high ploidy level, high polymorphism rate and aneuploidy, the introduction of CRISPR/Cas mutations are impaired. Many different factors can affect CRISPR/Cas mutation rate. One recently discovered is that a Cas enzyme construct containing introns from the target species can improve genome editing. Introns are capable of enhancing gene expression and mRNA accumulation via a process known as "Intron-mediated enhancement" or IME. However, the stimulation of the expression can be affected by other levels of regulation such as the formation of stable secondary structures that can downregulate gene expression, and alternative splicing. Based on this, this project aims to computationally explore pre-defined introns with IME enhancing properties and splice site facilitated recognition indicators using deep learning and thermodynamic approaches to predict secondary structure formation and stability to select sugarcane introns that do not likely form stable secondary structures and could thus theoretically enhance CRISPR/Cas mutation rate. (AU)