Transgenic maize: methods and protocols

Abstract

This book is number 526 in the popular ‘Methods in Molecular Biology’ series, which is renowned for its targeted protocol chapters, each with photographs, recipes and a set of notes for additional practical guidance and to avoid common pitfalls. It focuses specifically on domesticated maize (corn), which has come a long way from its teosinte origins in Mexico, to become one of the world's most important foods and feed crops, with a small but significant role in bio-fuel production. What makes this book particularly timely is that maize is now increasingly marketed as F1 hybrid seed in ‘biotech form’ with almost one-third of all maize sown globally in 2010 possessing genetically modified traits of insect resistance, herbicide tolerance, or both. This reference book takes the logical route of a typical research and development project, with chapters on: design of gene constructs; various transformation methods; use of transgenic maize in research; methods to analyse transgenic plants; and, finally, breeding with transgenics. There are useful tips on codon optimisation and the incorporation of 5′ UTRs, matrix attachment or scaffold-associated regions (MARS and SARS) and introns into transformation cassettes for maximising expression levels. The sections on fluorescent markers to localise translational fusion proteins, and RNAi to silence pre-defined maize genes are particularly interesting for scientists using transgenic approaches in their research. The four chapters on transformation methods compare biolistics, Agrobacterium tumefaciens and silicon carbide whiskers for DNA-delivery into immature zygotic embryos or competent callus cultures derived from HiII, H99 and other regenerable genotypes. They also provide detailed instructions for making and sterilising the various tissue culture media required and the commonly used selection systems based on resistance to specific antibiotics or herbicides. The detailed methods and additional notes make the transformation of maize accessible to anyone with a reasonably equipped laboratory and plant growth facilities. I particularly like the note in the chapter on DNA hybridisation that suggests ‘an unwanted book is ideal for weighing down the blotting stack’ and immediately pictured the author's stained PhD thesis faithfully holding down the absorbent towels. The analysis of transgenic plants includes chapters on: plasmid rescue as a method to recover flanking genomic sequences from gene-tagged lines; strategies for detecting rare sequences using non-radioactive DNA blots; high-throughput tissue printing of seeds; and the determination of transgene copy number by real-time PCR. The final chapter describes backcross breeding strategies used to introgress transgene events that are normally inherited as dominant loci into elite, adapted germplasm for commercialisation. There is a lot to like about this collection of practical chapters, which represent an excellent investment for the applied maize researcher, who could be a student doing an undergraduate project through to a professional scientist in a dedicated transformation laboratory. Although the best learning outcomes usually result from hands-on, actually-doing-it-yourself training, this book comes a close second best. There is a functional, expedient style to the writing, due in part to the fact that almost half the 21 contributing authors work for major biotechnology companies, with the majority of the others doing applied research at major US universities. My only gripes are the positioning of all colour plates together at the centre of the book, and the laughable index that lists only one or two topics per letter heading and could find nothing at all to reference against D, F, J, N or O!