CRISPR-Cas9 delivery via carbon nanotubes and chitosan nanoparticles for floral induction in Norway spruce
Main PI: Ove Nilsson, Department of Forest Genetics and Plant Physiology, SLU
Co-PI: Ulrika Egertsdotter, Department of Forest Genetics and Plant Physiology, SLU and Renewable Bioproducts Institute, Georgia Tech, Atlanta, GA, USA
Abstract
This project aims to identify genes controlling cone-setting to trigger early “flowering” in Picea abies to circumvent its long juvenile phase that hampers breeding and implementation of breeding efforts. The approach is to test and identify novel biotechnology tools for improvement of Picea abies by using carbon nanotubes for RNA delivery and to further explore the potential for gene editing utilizing chitosan nanoparticles for CRISPR-Cas9 constructs delivery into in vitro cultures of early-stage somatic embryos (proembryogenic masses, PEMs) of P. abies.
Objectives
1. Develop a protocol for RNA transfection of P. abies PEMs based on carbon nanotubes:
a.Design and have nanocarbon tubes (NCT) manufactured.
b.Functionalize NCT for transfection.
c.Test and optimize transfection process steps with the GFP reporter.
2.Explore the use of chitosan nanoparticles (CSNPs) for transfection in combination with NCT or alone.
3.Test and verify delivery of CRISPR-Cas9 ribonucleoprotein (RNP) complexes and single-guide RNAs (sgRNAs) directly into target tissues of P. abies using the optimal nanoparticle method determined in 1. and 2.
About the PIs and their synergies
Prof. Ove Nilsson is an expert on tree biotechnology and the molecular regulation of flowering. He has long experience with the use of CRISPR-Cas9 genome editing in trees. Ulrika Egertsdotter is an expert in spruce somatic embryogenesis and conifer transformation. This research will enable exploration of novel, non-GMO methods to generate trees with improved traits such as early flowering. The targeted area requires synergistic efforts based on in depth understanding of molecular biology and the physiology of the targeted plant material to identify a successful experimental approach. New insights from this project will benefit future goals within forestry to optimize breeding for production and climate adaptation.
