We report the construction of an RFLP genetic map of rice (Oryza sativa) chromosomes. The map is comprised of 135 loci corresponding to clones selected from a PstI genomic library. This molecular map covers 1,389 cM of the rice genome and exceeds the current classical maps by more than 20%. The map was generated from F2 segregation data (50 individuals) from a cross between an indica and javanica rice cultivar. Primary trisomics were used to assign linkage groups to each of the 12 rice chromosomes. Seventy-eight percent of the clones assayed revealed RFLPs between the two parental cultivars, indicating that rice contains a significant amount of RFLP variation. Strong correlations between size of hybridizing restriction fragments and level of polymorphism indicate that a significant proportion of the RFLPs in rice are generated by insertions/delections. This conclusion is supported by the occurrence of null alleles for some clones (presumably created by insertion or deletion events). One clone, RG229, hybridized to sequences in both the indica and javanica genomes, which have apparently transposed since the divergence of the two cultivars from their last common ancestor, providing evidence for sequence movement in rice. As a by product of this mapping project, we have discovered that rice DNA is less C-methylated than tomato or maize DNA. Our results also suggest the notion that a large fraction of the rice genome (approximately 50%) is single copy.

SUMMARY Mitogen-activated protein kinase cascades are important signaling modules that convert environmental stimuli into cellular responses. We show that MPK3, MPK4, and MPK6 are rapidly activated after cold treatment. The mpk3 and mpk6 mutants display increased expression of CBF genes and enhanced freezing tolerance, whereas constitutive activation of the MKK4/5-MPK3/6 cascade in plants causes reduced expression of CBF genes and hypersensitivity to freezing, suggesting that the MKK4/5-MPK3/6 cascade negatively regulates the cold response. MPK3 and MPK6 can phosphorylate ICE1, a bHLH transcription factor that regulates the expression of CBF genes, and the phosphorylation promotes the degradation of ICE1. Interestingly, the MEKK1-MKK2-MPK4 pathway constitutively suppresses MPK3 and MPK6 activities, and has a positive role in the cold response. Furthermore, the MAPKKK YDA, and two calcium/calmodulin-regulated receptor-like kinases, CRLK1 and CRLK2, negatively modulate the cold-activation of MPK3/6. Our results uncover important roles of MAPK cascades in the regulation of plant cold response.

The perception and relay of cell-wall signals are critical for plants to regulate growth and stress responses, but the underlying mechanisms are poorly understood. We found that the cell-wall leucine-rich repeat extensins (LRX) 3/4/5 are critical for plant salt tolerance in Arabidopsis. The LRXs physically associate with the RAPID ALKALINIZATION FACTOR (RALF) peptides RALF22/23, which in turn interact with the plasma membrane-localized receptor-like protein kinase FERONIA (FER). The lrx345 triple mutant as well as fer mutant plants display retarded growth and salt hypersensitivity, which are mimicked by overexpression of RALF22/23. Salt stress promotes S1P protease-dependent release of mature RALF22 peptides. Treatment of roots with mature RALF22/23 peptides or salt stress causes the internalization of FER. Our results suggest that the LRXs, RALFs, and FER function as a module to transduce cell-wall signals to regulate plant growth and salt stress tolerance.

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A set of 219 DNA clones derived from mungbean (Vigna radiata), cowpea (V. unguiculata), common bean (Phaseolus vulgaris), and soybean (Glycine max) were used to generate comparative linkage maps among mungbean, common bean, and soybean. The maps allowed an assessment of linkage conservation and collinearity among the three genomes. Mungbean and common bean, both of the subtribe Phaseolinae, exhibited a high degree of linkage conservation and preservation of marker order. Most linkage groups of mungbean consisted of only one or two linkage blocks from common bean (and vice versa). The situation was significantly different with soybean, a member of the subtribe Glycininae. Mungbean and common bean linkage groups were generally mosaics of short soybean linkage blocks, each only a few centimorgans in length. These results suggest that it would be fruitful to join maps of mungbean and common bean, while knowledge of conserved genomic blocks would be useful in increasing marker density in specific genomic regions for all three genera. These comparative maps may also contribute to enhanced understanding of legume evolution.

Both Pi-2(t) and Pi-4(t) genes of rice confer complete resistance to the blast fungal pathogen Pyricularia oryzae Cav. As economically important plant genes, they have been recently characterized phenotypically, yet nothing is known about their classical linkage associations and gene products. We report here the isolation of DNA markers closely linked to these blast resistance genes in rice. The DNA markers were identified by testing 142 mapped rice genomic clones as hybridization probes against Southern blots, consisting of DNA from pairs of nearly isogenic lines (NILs) with or without the target genes. Chromosomal segments introgressed from donor genomes were distinguished by restriction fragment length polymorphisms (RFLPs) between the NILs. Linkage associations of the clones with Pi-2(t) and Pi4(t) were verified using F3 segregating populations of known blast reaction. Cosegregation of the resistant genotype and donor-derived allele indicated the presence of linkage between the DNA marker and a blast resistance gene. RFLP analysis showed that Pi-2(t) is closely linked to a single-copy DNA clone RG64 on chromosome 6, with a distance of 2.8+1.4(SE) cMorgans. Another blast resistance gene, Pi-4(t), is 15.3+4.2(SE) cMorgans away from a DNA clone RG869 on chromosome 12. These chromosomal regions can now be examined with additional markers to define the precise locations of Pi-2(t) and Pi-4(t). Tightly linked DNA markers may facilitate early selection for blast resistance genes in breeding programs. These markers may also be useful to map new genes for resistance to blast isolates. They may ultimately lead to the cloning of those genes via chromosome walking. The gene tagging approach demonstrated in this paper may apply to other genes of interest for both monogenic and polygenic traits.

Decline disease causes severe damage to bayberry. However, the cause of this disease remains unclear. Interestingly, our previous studies found that the disease severity is related with the level of soil fertilizer. This study aims to explore the effect and mechanism of compound fertilizer (CF) and bio-organic fertilizer (OF) in this disease by investigating the vegetative growth, fruit characters, soil property, rhizosphere microflora and metabolites. Results indicated that compared with the disease control, CF and OF exhibited differential effect in plant healthy and soil quality, together with the increase in relative abundance of Burkholderia and Mortierella, and the reduction in that of Rhizomicrobium and Acidibacter, Trichoderma, and Cladophialophora reduced. The relative abundance of Geminibasidium were increased by CF (251.79%) but reduced by OF (13.99%). In general, the composition of bacterial and fungal communities in rhizosphere soil was affected significantly at genus level by exchangeable calcium, available phosphorus, and exchangeable magnesium, while the former two variables had a greater influence in bacterial communities than fungal communities. Analysis of GC-MS metabonomics indicated that compared to the disease control, CF and OF significantly changed the contents of 31 and 45 metabolites, respectively, while both fertilizers changed C5-branched dibasic acid, galactose, and pyrimidine metabolic pathway. Furthermore, a significant correlation was observed at the phylum, order and genus levels between microbial groups and secondary metabolites of bayberry rhizosphere soil. In summary, the results provide a new way for rejuvenation of this diseased bayberry trees.

Decline disease causes serious damage and rapid death in bayberry, an important fruit tree in south China, but the cause of this disease remains unclear. The aim of this study was to investigate soil quality, microbial community structure and metabolites of rhizosphere soil samples from healthy and diseased trees. The results revealed a significant difference between healthy and diseased bayberry in soil properties, microbial community structure and metabolites. Indeed, the decline disease caused a 78.24% and 78.98% increase in Rhizomicrobium and Cladophialophora, but a 28.60%, 57.18%, 38.84% and 68.25% reduction in Acidothermus, Mortierella, Trichoderma and Geminibasidium, respectively, compared with healthy trees, based on 16S and ITS amplicon sequencing of soil microflora. Furthermore, redundancy discriminant analysis of microbial communities and soil properties indicated that the main variables of bacterial and fungal communities included pH, organic matter, magnesium, available phosphorus, nitrogen and calcium, which exhibited a greater influence in bacterial communities than in fungal communities. In addition, there was a high correlation between the changes in microbial community structure and secondary metabolites. Indeed, GC–MS metabolomics analysis showed that the healthy and diseased samples differed over six metabolic pathways, including thiamine metabolism, phenylalanine–tyrosine–tryptophan biosynthesis, valine–leucine–isoleucine biosynthesis, phenylalanine metabolism, fatty acid biosynthesis and fatty acid metabolism, where the diseased samples showed a 234.67% and 1007.80% increase in palatinitol and cytidine, respectively, and a 17.37%–8.74% reduction in the other 40 metabolites compared to the healthy samples. Overall, these results revealed significant changes caused by decline disease in the chemical properties, microbiota and secondary metabolites of the rhizosphere soils, which provide new insights for understanding the cause of this bayberry disease.