Commercial cultivars of garlic, a popular condiment, are sterile, making genetic studies and breeding of this plant challenging. However, recent fertility restoration has enabled advanced physiological and genetic research and hybridization in this important crop. Morphophysiological studies, combined with transcriptome and proteome analyses and quantitative PCR validation, enabled the identification of genes and specific processes involved in gametogenesis in fertile and male-sterile garlic genotypes. Both genotypes exhibit normal meiosis at early stages of anther development, but in the male-sterile plants, tapetal hypertrophy after microspore release leads to pollen degeneration. Transcriptome analysis and global gene-expression profiling showed that >16,000 genes are differentially expressed in the fertile vs. male-sterile developing flowers. Proteome analysis and quantitative comparison of 2D-gel protein maps revealed 36 significantly different protein spots, 9 of which were present only in the male-sterile genotype. Bioinformatic and quantitative PCR validation of 10 candidate genes exhibited significant expression differences between male-sterile and fertile flowers. A comparison of morphophysiological and molecular traits of fertile and male-sterile garlic flowers suggests that respiratory restrictions and/or non-regulated programmed cell death of the tapetum can lead to energy deficiency and consequent pollen abortion. Potential molecular markers for male fertility and sterility in garlic are proposed.

Garlic (Allium sativum L.) cultivars do not develop fertile flowers and seeds. Therefore, garlic production and improvement depend exclusively on vegetative propagation. Recent advances in garlic research have enabled fertility restoration and the discovery of fertile and male-sterile genotypes; however, the environmental regulation of the reproductive process is still not clear. Garlic seeds are successfully produced in the Mediterrenean region, where the photoperiod is relatively short, whereas spring and summer temperatures are high. We hypothesise that, in bolting garlic, various stages of florogenesis are differentially regulated by temperature and that high temperatures might obstruct pollen production. The effects of eight combinations of controlled growth temperatures on fertile and male-sterile garlic clones were studied. In both genotypes, a gradual temperature increase before and during anthesis favoured intact flower development. Surprisingly, continuous exposure to moderate temperatures during the entire growth period resulted in poor flowering, anther abortion and reduced pollen production. In the male-sterile genotype, no growth regime improved pollen production, which is controlled by genetic mechanisms. In the male-fertile genotype, gradual temperature increase supported pollen production but a sharp transition to high temperatures resulted in rapid flower senescence and pollen abortion, thus supporting our research hypothesis. In both fertile and male-sterile plants, the most vulnerable phase of microsporogenesis is the unicellular microspore stage. Tapetal malformation is the major cause for malnutrition of the microspores, with consequent production of nonviable pollen grains.

Background Commercial cultivars of garlic produce neither flowers nor seeds, and therefore efficient breeding of this important vegetable and medicinal crop is impossible. New avenues for hybridization and seed production were recently opened by the restoration of garlic fertility. The huge genome of garlic was fully sequenced in 2020 but, before that, transcriptome catalogs had been employed as a basis for garlic genetic studies. Here, we provide the conjoint genomic and transcriptome analysis of the regulatory network in flowering garlic genotypes. Results Our genome-wide analysis revealed phosphatidylethanolamine-binding proteins (PEBP) and LEAFY (LFY) genes that were not previously observed at the transcriptome level. Functions of TFL-like genes were reduced and probably replaced by FT-like homologs, whereas homologs of MFT-like genes were not found in the garlic genome. The discovery of three sequences of LFY-like genes in the garlic genome and confirmation of their alternative splicing suggest their key role in garlic florogenesis. It is not yet clear whether only one gene—AsLFY1—acts alone as the "pioneer transcription factor" or AsLFY2 also provides these functions. Conclusions The garlic genome, including key flowering genes, is highly repetitive. The presence of several orthologs of flowering genes that differ in their temporal and spatial expression and co-expression network advocates ongoing evolution in the garlic genome and diversification of gene functions. We propose that the process of fertility deprivation in garlic cultivars is based on the loss of transcriptional functions of the specific genes. Further comparisons of the genome and transcriptome factors in flowering and non-flowering garlic genotypes will clarify the possible evolution of their reproductive traits and contribute to fertility restoration and efficient breeding of this important crop.