Epsilon Archive for Student Projects

Abstract

A growing world population, a changing climate with changed agricultural conditions, and animal welfare are a few reasons behind the need for a more sustainable and local food chain. Pea (Pisum sativum L.) has been globally recognised as a candidate for a green protein with several health benefits and a high nutritional value. Chemical compounds called anti-nutrients, such as phytic acid, lectins, and saponins, affect the health benefits by compromising the uptake of nutrients. The Grogrund research project “Pea – the key for future green protein” at the Swedish University of Agricultural Sciences (SLU) aims to improve the nutritional profile of the two commercial cultivars Clara and Ingrid by reducing the amount of anti-nutritional factors using CRISPR. This thesis is part of that project, and the goal is to start the development of a protocol for tissue culture for the two pea cultivars. Tissue culture is an important step in plant breeding, as it serves as a tool for producing a whole new plant with CRISPR-induced characteristics. Developing a protocol is a time-consuming process. Pea is not very responsive to tissue culture with a low regeneration rate, and the protocol needs to be adjusted after each specific cultivar. The best approach is to start with already existing protocols and then keep on working with the most promising ones. Ten different protocols for callus induction and shoot induction were tested, as well as a protocol for root induction. Six different explants were tried: cotyledons cut in three different ways, epicotyls, hypocotyls, and leaves. The hormones used were BAP, NAA, 2,4-D, TDZ, kinetin, IAA, and IBA. A stable protocol has a high regeneration rate and ensures that the new genetic traits are present throughout the whole plant and passed on to the next generations. The results from this study show that the cultivars react very differently to the ten treatments. Cotyledons and hypocotyls developed the highest number of shoots, and the results indicate that two of the ten media seem promising. The one with 0.5 mg L-1 BAP and 4.0 m L-1 NAA for callus induction media, and 1.0 mg L-1 BAP and 0.25 mg L-1 NAA for shoot induction media, and the one with 2.0 mg L-1 BAP and 2.0 mg L-1 NAA for callus induction media and 4.5 mg L-1 BAP and 0.2 mg L-1 NAA for shoot induction media.