A CYC/TB1-type TCP transcription factor controls spikelet meristem identity in barley.

Barley possesses a branchless, spike-shaped inflorescence where determinate spikelets attach directly to the main axis, but the developmental mechanism of spikelet identity remains largely unknown. Here we report the functional analysis of the barley gene BRANCHED AND INDETERMINATE SPIKELET 1 (BDI1), which encodes a TCP transcription factor and plays a crucial role in determining barley inflorescence architecture and spikelet development. The bdi1 mutant exhibited indeterminate spikelet meristems that continued to grow and differentiate after producing a floret meristem; some spikelet meristems at the base of the spike formed two fully developed seeds or converted to branched spikelets, producing a branched inflorescence. Map-based cloning analysis showed that this mutant has a deletion of ~600 kb on chromosome 5H containing three putative genes. Expression analysis and virus-induced gene silencing confirmed that the causative gene, BDI1, encodes a CYC/TB1-type TCP transcription factor and is highly conserved in both wild and cultivated barley. Transcriptome and regulatory network analysis demonstrated that BDI1 may integrate regulation of gene transcription cell wall modification and known trehalose-6-phosphate homeostasis to control spikelet development. Together, our findings reveal that BDI1 represents a key regulator of inflorescence architecture and meristem determinacy in cereal crop plants.

Physical organization of repetitive sequences and chromosome diversity of barley revealed by fluorescence in situ hybridization (FISH).

Fluorescence in situ hybridization (FISH) using oligonucleotides is a simple and convenient method for chromosome research. In this study, 34 of 46 previously developed oligonucleotides produced signals in barley. Together with two plasmid clones and one PCR-amplified cereal centromere repeat (CCS1) probe, 37 repetitive sequences were chromosomally located produced three types of signals covering different positions on the chromosomes. The centromeric and pericentric regions had a more complex genomic organization and sequence composition probably indicative of higher contents of heterochromatin. An efficient multi-plex probe containing eight oligonucleotides and a plasmid clone of 45S rDNA was developed. Thirty-three barley karyotypes were developed and compared. Among them, 11 irradiation-induced mutants of cultivar 08-49 showed no chromosomal variation, whereas 22 cultivar and landrace accessions contained 28 chromosomal polymorphisms. Chromosome 4H was the most variable and 6H was the least variable based on chromosome polymorphic information content (CPIC). Five polymorphic chromosomes (1H-2, 2H-1, 3H-3, 5H-2, and 6H-2) were dominant types, each occurring in more than 50% of accessions. The multi-plex probe should facilitate identification of further chromosomal polymorphisms in barley.

Structural chromosome rearrangements and polymorphisms identified in Chinese wheat cultivars by high-resolution multiplex oligonucleotide FISH.

KEY MESSAGE: High-resolution multiplex oligonucleotide FISH revealed the frequent occurrence of structural chromosomal rearrangements and polymorphisms in widely grown wheat cultivars and their founders. Over 2000 wheat cultivars including 19 founders were released and grown in China from 1949 to 2000. To understand the impact of breeding selection on chromosome structural variations, high-resolution karyotypes of Chinese Spring (CS) and 373 Chinese cultivars were developed and compared by FISH (fluorescence in situ hybridization) using an oligonucleotide multiplex probe based on repeat sequences. Among them, 148 (39.7%) accessions carried 14 structural rearrangements including three single translocations (designated as T), eight reciprocal translocations (RT), one pericentric inversion (perInv), and two combined variations having both the deletion and single translocations. Five rearrangements were traced to eight founders, including perInv 6B detected in 57 cultivars originating from Funo, Abbondanza, and Fan 6, T 1RS∙1BL in 47 cultivars derived from the Lovrin series, RT 4AS∙4AL-1DS/1DL∙1DS-4AL in 31 varieties from Mazhamai and Bima 4, RT 1RS∙7DL/7DS∙1BL in three cultivars was from Aimengniu, and RT 5BS∙5BL-5DL/5DS∙5DL-5BL was only detected in Youzimai. In addition to structural rearrangements, 167 polymorphic chromosome blocks (defined as unique signal patterns of oligonucleotide repeat probes distributed within chromosomes) were identified, and 59 were present in one or more founders. Some specific types were present at high frequencies indicating selective blocks in Chinese wheat varieties. All cultivars and CS were clustered into four groups and 15 subgroups at chromosome level. Common block patterns occurred in the same subgroup. Origin, geographic distribution, probable adaptation to specific environments, and potential use of these chromosomal rearrangements and blocks are discussed.

A simple and efficient non-denaturing FISH method for maize chromosome differentiation using single-strand oligonucleotide probes.

Single-strand oligonucleotides (SSONs hereafter) as probes are becoming a powerful method of chromosome painting in many species. In this study, nine SSONs ((ACT)10, (ACT)19, Knob-1, Knob-2, Knob-3, CentC69-1, MR68-3, K10-72-1, and TR1-357-2) were developed and used for chromosome identification in 16 maize (Zea mays L., 2n = 20) inbred lines and hybrids by non-denaturing fluorescence in situ hybridization (ND-FISH). Each SSON produced clear signals on 2-10 chromosomes of inbred lines B73 and Mo17. A multiplex probe set containing four SSONs ((ACT)10, Knob-2, CentC69-1, and MR68-3) clearly characterized all maize chromosomes in the 16 lines by a single round of ND-FISH and revealed genetic variation at a chromosome level. For example, unique signals on chromosome 6 clearly distinguished all 16 genotypes. The SSONs and multiplex probe developed in this research will facilitate genotype identification and chromosome research in maize.