Meloidogyne incognita is a devastating plant-parasitic nematode.  Effectors play important roles during the stages of nematodes infection and parasitism, but their molecular functions remain largely unknown.  In this study, we characterized a new effector, Minc03329, which contains signal peptide for secretion and a C-type lectin domain.  The yeast signal sequence trap experiments indicated that the signal peptide of Minc03329 is functional.  In situ hybridization showed that Minc03329 was specifically expressed in the subventral esophageal gland.  Real-time qPCR confirmed that the expression level of Minc03329 transcript was significantly increased in pre-parasitic and parasitic second-stage juveniles (pre-J2s and par-J2s).  Tobacco rattle virus (TRV)-mediated gene silencing of Minc03329 in host plants largely reduced the pathogenicity of nematodes.  On the contrary, ectopic expression of Minc03329 in Arabidopsis thaliana significantly increased plant susceptibility to nematodes.  Transient expression of Minc03329 in Nicotiana benthamiana leaves suppressed the programmed cell death triggered by the pro-apoptotic protein BAX.  Moreover, the transcriptome analysis of Minc03329-transgenic Arabidopsis and wild type revealed that many defense-related genes were significantly down-regulated.  Interestingly, some different expressed genes were involved in the formation of nematode feeding sites.  These results revealed that Minc03329 is an important effector for M. incognita, suppressing host defense response and promoting pathogenicity.

The expression of mitogen-activated protein kinase (mapk) double-stranded RNA in cucumber is effective in controlling infestations of the root-knot nematode Meloidogyne incognita.  However, little is known about the ecological effects of transgenic plants.  Here, we analyzed the diversity of 16S rDNA genes derived from the rhizosphere archaea of transgenic cucumber plants as an indicator of ecological change.  A total of 17 and 18 operational taxonomic units were detected in the rhizospheres of non-transgenic cucumber and mapk dsRNA-expressing plants, respectively.  No significant difference was observed between the two groups according to Shannon and Simpson indices.  In soil samples of the two rhizospheres, the dominant group was Crenarchaeota at the phylum level, with Staphylothermus, Methanococcus, Pyrodictium and Sulfolobus the abundant taxa at the genus level.  These results suggest that expressing mapk double-stranded (ds) RNA in cucumber has no apparent effect on the diversity of rhizosphere archaea, and provide powerful evidence for the ecological safety of transgenic cucumber expressing mapk dsRNA.   

   Virus-tolerant plant, which allows the accumulation of virus and then generates virus-derived small RNAs (vsRNAs), is a valuable material to reveal the antiviral efficiency of vsRNAs. Here, a comparison of vsRNAs in Tomato yellow leaf curl virus tolerant and in susceptible tomato varieties showed the consistent trend of vsRNAs’ distribution on virus genome, which is presented as an obvious characteristic. However, the expression level of vsRNA in tolerant variety is less than that in susceptible variety. Slicing targets of vsRNA-mediated viral transcripts were investigated using parallel analysis of RNA ends, and geminivirus DNA methylation was determined by bisulfite sequencing, which uncovered that not all vsRNAs participated in viral mRNA degradation and DNA methylation. Additionally, by comparing with the expression pattern of vsRNAs, viral DNA and mRNA, we proposed the quantity of vsRNAs is corresponding to the expression level of viral mRNA, while the virus-suppression of vsRNAs is not high-efficient.

Fusarium oxysporum f. sp. conglutinans (Foc) is the causal agent of Fusarium wilt disease of Brassica oleracea. A rapid, accurate, and reliable method to detect and identify plant pathogens is vitally important to integrated disease management. In this study, using a comparative genome analysis among Fusarium oxysporum (Fo), we developed a Foc-specific primer set (Focs-1/Focs-2) and established a multiplex-PCR assay. In the assay, the Focs-1/Focs-2 and universal primers for Fusarium species (W106R/F106S) could be used to detect Foc isolates in a single PCR reaction. With the optimized PCR parameters, the multiplex-PCR assay showed a high specificity for detecting Foc and was very sensitive to detect as little as 100 pg of pure Foc genomic DNA or 1 000 spores in 1 g of twice-autoclaved soil. We also demonstrated that Foc isolates were easily detected from infected plant tissues, as well as from natural field soils, using the multiplex-PCR assay. To our knowledge, this is a first report on detection Fo by comparative genomic method.

Telomeres form the ends of eukaryotic chromosomes and serve as protective caps that keep chromosomes structure independency and completeness. The first plant telomere DNA was isolated from Arabidopsis thaliana and was shown to have tandemly repeated sequence 5´-TTTAGGG-3´. The Arabidopsis-type telomere has been found in many plants, but several reports indicate that this sequence is absent in some plants. Up to now, no research has been conducted on the telomere of cotton. In this paper, the Arabidopsis-type telomere sequence was amplified and cloned using the primers designed based on the fragment containing telomere sequence in an Arabidopsis bacterial artificial chromosome (BAC). Fluorescence in situ hybridization (FISH) with cotton metaphase chromosomes using the Arabidopsis-type telomere sequence as probes indicated that the signals were located at all chromosome ends of seven diploid and two tetraploid cotton species with different signal intensities among chromosome complements of different cotton species, even between long and short arms of the same chromosome. To identify the signals of FISH, the genome DNA of Xinhai 7, a cultivar of Gossypium barbadense, digested by BAL-31 nuclease was introduced in this study. The result of BAL-31 digestion indicated that the hybridization signals of FISH represent the outermost DNA sequence of each cotton chromosomes. So we first proved that the telomeric repeats of cotton cross-hybridize with that of Arabidopsis. The results of terminal restriction fragment (TRF) showed significant variation in telomere length among cotton species. The telomere length of cultivated cotton was close to 20 kb and was larger than those of wild cotton species whose telomere length ranged from 6 to 20 kb.

Red fruit peel is one of the most valuable economic traits in pear and is mainly determined by anthocyanins. Many pear cultivars with a red peel originated from bud sports; however, little is known about the genetic mechanisms underlying this trait. We have previously identified a mutant PpBBX24 containing a 14-nucleotide deletion in the coding region (Ppbbx24-del) as the only known variant associated with the red coloration of the mutant 'Red Zaosu' pear (Pyrus pyrifolia White Pear Group). Herein, we analyzed the role of the mutant gene in red coloration and its mechanism of action. The results showed that light promoted red peel coloration in 'Red Zaosu' pear, and Ppbbx24-del had positive effects on light-induced anthocyanin biosynthesis, while normal PpBBX24 had the opposite effects. Transient and stable transformation experiments confirmed that Ppbbx24-del could promote anthocyanin accumulation in pear fruit peels, calli, and tobacco flowers. Due to the loss of NLS and VP domains, Ppbbx24-del co-localized in the nucleus and cytoplasm, whereas PpBBX24 localized only in the nucleus. Real-time PCR and transcriptome analyses indicated that PpMYB10 and PpHY5 is highly expressed in 'Red Zaosu' pear. In yeast one-hybrid and dual luciferase assays, Ppbbx24-del and PpHY5 independently promoted the expression of PpCHS, PpCHI, and PpMYB10 by binding to their promoters; however, PpBBX24 did not affect the expression of these genes. Additionally, we found that Ppbbx24-del and PpHY5 had additive effects on the expression of PpCHS, PpCHI, and PpMYB10, as they promote the expression of anthocyanin synthesis genes separately. The co-expression of PpBBX24 and PpHY5 inhibited the activation of downstream genes by PpHY5, and this was attributed to the interaction between the two loci. In conclusion, our results clarify the molecular mechanism by which mutant Ppbbx24-del and PpBBX24 exert opposite effects in the regulation of anthocyanin accumulation in pear. These findings lay an important theoretical foundation for the use of Ppbbx24-del to create red pear cultivars.