Agrobacterium tumefaciens mediated plant transformation is a versatile tool for plant genetic engineering following its discovery nearly half a century ago. Numerous modifications were made in its application to increase efficiency, especially in the recalcitrant major cereals plants. Recent breakthroughs in transformation efficiency continue its role as a mainstream technique in CRISPR/Cas-based genome editing and gene stacking. These modifications led to higher transformation frequency and lower but more stable transgene copies with the capability to revolutionize modern agriculture. In this review, we provide a brief overview of the history of Agrobacterium-mediated plant transformation and focus on the most recent progress to improve the system in both the Agrobacterium and the host recipient. A promising future for transformation in biotechnology and agriculture is predicted.

Common wheat (Triticum aestivum) is a hexaploid plant (AABBDD) derived from genetically related tetraploid wheat T. turgidum (AABB) and a diploid goatgrass Aegilops tauschii (DD). Recent advances in sequencing technology and genome assembly strategies allow the acquisition of multiple wheat genomes, calling for a centralized database to store, manage and query the genomics information in a manner to reflect their evolutionary relationship and to perform effective comparative genome analysis. Here, we built WheatGene, a database that contains five wheat genomes of 318,102 genes and 945,900 transcripts and their expression information in 998 RNA-seq samples that can be searched and compared in an interactive manner. WheatGene was developed with Drupal, a popular content management system and the toolkit Tripal managed the biological information. The database was accessible through a web browser with species, search, gene expression, tools, and literature entries. Tools available were BLAST, synteny viewer, map viewer, JBrowse, data downloads, gene expression heatmap and bar chart, and homologs viewer. Moreover, the map viewer connected genomics data with genetic maps and QTL that can be searched for markers for molecular breeding. WheatGene was developed with open-source modules and libraries. WheatGene is available at http://wheatgene.agrinome.org.

The Q gene in common wheat encodes an APETALA2 (AP2) transcription factor that causes the free threshing attribute. Wheat spikelets bearing several florets are subtended by a pair of soft glumes that allow free liberation of seeds. In wild species, the glumes are tough and rigid, making threshing difficult. However, the nature of these “soft glumes”, caused by the domestication allele Q is not clear. Here, we found that over expression of Q in common wheat leads to homeotic florets at glume positions. We provide phenotypic, microscopy, and marker genes evidence to demonstrate that the soft glumes of common wheat are in fact lemma-like organs, or so-called sterile-lemmas. By comparing the structures subtending spikelets in wheat and other crops such as rice and maize, we found that AP2 genes may play conserved functions in grasses by manipulating vestigial structures, such as floret-derived soft glumes in wheat and empty glumes in rice. Conversion of these seemingly vegetative organs to reproductive organs may be useful in yield improvement of crop species.

Bread wheat is not only an important cereal crop but also a model for study of an allopolyploid plant with a large, highly repetitive genome. Advances in next-generation sequencing (NGS) technology provide needed throughput to conquer the enormous size of the wheat genome. Multiple high quality reference genome sequences will soon be available. Full-scale wheat functional genomics studies are dawning. In this review we highlight the available tools and methodologies for wheat functional genomics research developed with the assistance of NGS technology and recent progress, particularly the concerted effort in generating multiple reference genomes, strategies to attain genome-wide genetic variation, genome-wide association studies, mutant population generation, and NGS-supported gene cloning and functional characterization. These resources and platforms lay a solid foundation for wheat research, leading to a new era of wheat functional genomics that will bridge the gap between genotype and phenotype. Dissection of wheat genomes and gene functions should assist in genomics-assisted selection and facilitate breeding of elite varieties for sustainable agriculture in China and the world.

The CRISPR/Cas9 technology is evolved from a type II bacterial immune system and represents a new generation of targeted genome editing technology that can be applied to nearly all organisms. Site-specific modification is achieved by a single guide RNA (usually about 20 nucleotides) that is complementary to a target gene or locus and is anchored by a protospacer-adjacent motif. Cas9 nuclease then cleaves the targeted DNA to generate double-strand breaks (DSBs), which are subsequently repaired by non-homologous end joining (NHEJ) or homology-directed repair (HDR) mechanisms. NHEJ may introduce indels that cause frame shift mutations and hence the disruption of gene functions. When combined with double or multiplex guide RNA design, NHEJ may also introduce targeted chromosome deletions, whereas HDR can be engineered for target gene correction, gene replacement, and gene knock-in. In this review, we briefly survey the history of the CRISPR/Cas9 system invention and its genome-editing mechanism. We also describe the most recent innovation of the CRISPR/Cas9 technology, particularly the broad applications of modified Cas9 variants, and discuss the potential of this system for targeted genome editing and modification for crop improvement.

Hexaploid wheat has triplicated homoeologs for most of the genes that are located in subgenomes A, B, and D. GASR7, a member of the Snakin/GASA gene family, has been associated with grain length development in wheat. However, little is known about divergence of its homoeolog expression in wheat polyploids. We studied the expression patterns of the GASR7 homoeologs in immature seeds in a synthetic hexaploid wheat line whose kernels are slender like those of its maternal parent (Triticum turgidum, AABB, PI 94655) in contrast to the round seed shape of its paternal progenitor (Aegilops tauschii, DD, AS2404). We found that the B homoeolog of GASR7 was the main contributor to the total expression level of this gene in both the maternal tetraploid progenitor and the hexaploid progeny, whereas the expression levels of the A and D homoeologs were much lower. To understand possible mechanisms regulating different GASR7 homoeologs, we firstly analyzed the promoter sequences of three homoeologous genes and found that all of them contained gibberellic acid (GA) response elements, with the TaGASR7B promoter (pTaGASR7B) uniquely characterized by an additional predicted transcriptional enhancer. This was confirmed by the GA treatment of spikes where all three homoeologs were induced, with a much stronger response for TaGASR7B. McrBC enzyme assays showed that the methylation status at pTaGASR7D was increased during allohexaploidization, consistent with the repressed expression of TaGASR7D. For pTaGASR7A, the distribution of repetitive sequence-derived 24-nucleotide (nt) small interfering RNAs (siRNAs) were found which suggests possible epigenetic regulation because 24-nt siRNAs are known to mediate RNA-dependent DNA methylation. Our results thus indicate that both genetic and epigenetic mechanisms may be involved in the divergence of GASR7 homoeolog expression in polyploid wheat.