Although long-term genetic gain has been achieved through increasing use of modern breeding methods and technologies, the rate of genetic gain needs to be accelerated to meet humanity's demand for agricultural products. In this regard, genomic selection (GS) has been considered most promising for genetic improvement of the complex traits controlled by many genes each with minor effects. Livestock scientists pioneered GS application largely due to livestock's significantly higher individual values and the greater reduction in generation interval that can be achieved in GS. Large-scale application of GS in plants can be achieved by refining field management to improve heritability estimation and prediction accuracy and developing optimum GS models with the consideration of genotype-by-environment interaction and non-additive effects, along with significant cost reduction. Moreover, it would be more effective to integrate GS with other breeding tools and platforms for accelerating the breeding process and thereby further enhancing genetic gain. In addition, establishing an open-source breeding network and developing transdisciplinary approaches would be essential in enhancing breeding efficiency for small- and medium-sized enterprises and agricultural research systems in developing countries. New strategies centered on GS for enhancing genetic gain need to be developed.

Enhancing Genetic Gain through Genomic Selection: From Livestock to Plants.

Sorghum is the fifth most widely planted cereal crop in the world and is commonly cultivated in arid and semi-arid regions such as Africa. Despite its importance as a food source, sorghum genetic improvement through transgenic approaches has been limited because of an inefficient transformation system. Here, we report a ternary vector (also known as cohabitating vector) system using a recently described pVIR accessory plasmid that facilitates efficient Agrobacterium-mediated transformation of sorghum. We report regeneration frequencies ranging from 6% to 29% in Tx430 using different selectable markers and single copy, backbone free 'quality events' ranging from 45% to 66% of the total events produced. Furthermore, we successfully applied this ternary system to develop transformation protocols for popular but recalcitrant African varieties including Macia, Malisor 84-7 and Tegemeo. In addition, we report the use of this technology to develop the first stable CRISPR/Cas9-mediated gene knockouts in Tx430.

https://onlinelibrary.wiley.com/doi/pdf/10.1111/pbi.12879

Developing a flexible, high-efficiency Agrobacterium-mediated sorghum transformation system with broad application.

Agriculture is an important source of human food. However, current agricultural practices need modernizing and strengthening to fulfill the increasing food requirements of the growing worldwide population. Genome editing (GE) technology has been used to produce plants with improved yields and nutritional value as well as with higher resilience to herbicides, insects, and diseases. Several GE tools have been developed recently, including clustered regularly interspaced short palindromic repeats (CRISPR) with nucleases, a customizable and successful method. The main steps of the GE process involve introducing transgenes or CRISPR into plants via specific gene delivery systems. However, GE tools have certain limitations, including time-consuming and complicated protocols, potential tissue damage, DNA incorporation in the host genome, and low transformation efficiency. To overcome these issues, nanotechnology has emerged as a groundbreaking and modern technique. Nanoparticle-mediated gene delivery is superior to conventional biomolecular approaches because it enhances the transformation efficiency for both temporal (transient) and permanent (stable) genetic modifications in various plant species. However, with the discoveries of various advanced technologies, certain challenges in developing a short-term breeding strategy in plants remain. Thus, in this review, nanobased delivery systems and plant genetic engineering challenges are discussed in detail. Moreover, we have suggested an effective method to hasten crop improvement programs by combining current technologies, such as speed breeding and CRISPR/Cas, with nanotechnology. The overall aim of this review is to provide a detailed overview of nanotechnology-based CRISPR techniques for plant transformation and suggest applications for possible crop enhancement.

/pdf/advantage-of-nanotechnology-based-genome-editing-system-and-41yjihu3gb.pdf

Advantage of Nanotechnology-Based Genome Editing System and Its Application in Crop Improvement.

The use of Baby boom (Bbm) and Wuschel2 (Wus2) has made maize transformation more efficient across an increasingly wide range of inbreds. However, the benefits have come with the requirement of excising these transformation helper components to enable plant regeneration, which adds size to the T-DNA, and complexity to the transformation system. A new system with the advantages of smaller size and simplicity for the selectable marker gene-containing T-DNA is described. First, expression of Zm-Wus2 alone driven by the maize Pltp promoter (Zm-Pltppro), was determined to be sufficient to induce rapid somatic embryo formation from the scutella of maize immature embryos. It was also demonstrated that co-infecting with two strains of Agrobacterium, one with a Wus2 expression cassette, and the other with a combination of both selectable and visual marker cassettes, produced transformed T0 plants that contained only a single copy of the selectable marker T-DNA, without the integration of Wus2. Furthermore, the process was optimized by varying the ratio of the two Agrobacterium strains, and by modulating Wus2 expression to enable high-frequency recovery of selectable marker-containing T0 plants that did not contain Wus2. Several factors may have contributed to this outcome. Wus2 expression in localized cell(s) appeared to stimulate somatic embryogenesis in neighboring cells, including those that had integrated the selectable marker. In addition, in cells in which the Wus2 T-DNA did not integrate but the selectable marker T-DNA did, transient Wus2 expression stimulated somatic embryo formation and regeneration of stable T0 plants that contained the selectable marker. In addition, augmenting the Pltp promoter with three viral enhancer elements to increase Wus2 expression stimulated embryogenesis while precluding their regeneration. The phenomenon has now been designated as “altruistic transformation.”

/pdf/use-of-non-integrating-zm-wus2-vectors-to-enhance-maize-2cnr6s5252.pdf

Use of non-integrating Zm-Wus2 vectors to enhance maize transformation

Edit at will: Genotype independent plant transformation in the era of advanced genomics and genome editing.

A simple and versatile ternary vector system that utilizes improved accessory plasmids for rapid maize transformation is described. This system facilitates high-throughput vector construction and plant transformation. The super binary plasmid pSB1 is a mainstay of maize transformation. However, the large size of the base vector makes it challenging to clone, the process of co-integration is cumbersome and inefficient, and some Agrobacterium strains are known to give rise to spontaneous mutants resistant to tetracycline. These limitations present substantial barriers to high throughput vector construction. Here we describe a smaller, simpler and versatile ternary vector system for maize transformation that utilizes improved accessory plasmids requiring no co-integration step. In addition, the newly described accessory plasmids have restored virulence genes found to be defective in pSB1, as well as added virulence genes. Testing of different configurations of the accessory plasmids in combination with T-DNA binary vector as ternary vectors nearly doubles both the raw transformation frequency and the number of transformation events of usable quality in difficult-to-transform maize inbreds. The newly described ternary vectors enabled the development of a rapid maize transformation method for elite inbreds. This vector system facilitated screening different origins of replication on the accessory plasmid and T-DNA vector, and four combinations were identified that have high (86–103%) raw transformation frequency in an elite maize inbred.

/pdf/an-improved-ternary-vector-system-for-agrobacterium-mediated-49qflpiy4i.pdf

An improved ternary vector system for Agrobacterium-mediated rapid maize transformation.

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An improved ternary vector system for Agrobacterium-mediated rapid maize transformation.