The PDZ-GEF Gef26 regulates synapse development and function via FasII and Rap1 at the Drosophila neuromuscular junction

Guanine nucleotide exchange factors (GEFs) are essential for small G proteins to activate their downstream signaling pathways, which are involved in morphogenesis, cell adhesion, and migration. Mutants of Gef26, a PDZ-GEF (PDZ domain-containing guanine nucleotide exchange factor) in Drosophila, exhibit strong defects in wings, eyes, and the reproductive and nervous systems. However, the precise roles of Gef26 in development remain unclear. In the present study, we analyzed the role of Gef26 in synaptic development and function. We identified significant decreases in bouton number and branch length at larval neuromuscular junctions (NMJs) in Gef26 mutants, and these defects were fully rescued by restoring Gef26 expression, indicating that Gef26 plays an important role in NMJ morphogenesis. In addition to the observed defects in NMJ morphology, electrophysiological analyses revealed functional defects at NMJs, and locomotor deficiency appeared in Gef26 mutant larvae. Furthermore, Gef26 regulated NMJ morphogenesis by regulating the level of synaptic Fasciclin II (FasII), a well-studied cell adhesion molecule that functions in NMJ development and remodeling. Finally, our data demonstrate that Gef26-specific small G protein Rap1 worked downstream of Gef26 to regulate the level of FasII at NMJs, possibly through a βPS integrin-mediated signaling pathway. Taken together, our findings define a novel role of Gef26 in regulating NMJ development and function.     

马铃薯实生群体遗传多样性的SSR分析

以云南马铃薯品种‘剑川红’植株1个浆果中的天然实生种子产生的70个单株以及B20[CIP010(♀)×CIP004(♂)]杂交组合后代100个实生苗单株为材料,采用12对SSR引物对自交种和杂交实生群体的遗传差异性进行分析,旨在从后代群体中找到与母本(亲本)在分子水平上表现一致的植株,为种质资源的长期保存提供依据。结果表明:(1)‘剑川红’自交群体的多态性比率为81.6%,比杂交组合B20实生群体的多态性比率72.8%略高,说明2个群体的多态性比率均较高。(2)聚类分析结果显示,自交后代和杂交后代群体的遗传相似系数均较高,变化范围均在0.74~0.96之间,说明2个群体均发生了不同程度的遗传分离,但分离的程度较小,绝大多数条带表现一致。(3)在所有供试材料中,同一浆果中均未发现与‘剑川红’母株在分子水平上表现完全一致的单株。研究认为,在分子水平上寻找完全不分离的实生群体难度非常大,需进一步评价与母株(亲本)在分子水平上相似株系的田间表现,从而确定是否可以通过相近或极相近株系来恢复种源。     

An improved semiautomated rapid method of extracting genomic DNA for molecular marker analysis in cocoa,<Emphasis Type="Italic">Theobroma cacao</Emphasis> L

DNA extraction is a time-consuming and expensive component of molecular marker analysis, constituting about 30–60% of the total time required for sample processing. Furthermore, the procedure for extracting high-quality DNA from tree species such as cocoa differs from extraction protocols suitable for other crop plants. This is accompanied by problems in collecting leaf tissues from field-grown cocoa trees, where storage facilities are not available and where transporting samples to laboratory for immediate refrigeration is usually impossible. We preserved cocoa leaf tissues in the field in an NaCl-CTAB-azide solution (as described in Rogstad, 1992), which did not require immediate refrigeration. This method also allowed preservation of leaf tissues for a few days during transportation and protected leaf tissues from bacterial and fungal attacks. Once transported to the laboratory, the samples were stored at 4°C for almost 1 y. To isolate good-quality DNA from stored leaf tissues, a rapid semiautomated and relatively high-throughput protocol was established. The procedure followed a modified CTAB/β-mercaptoethanol method of DNA extraction in a 96-well plate, and an automated system (i.e., GenoGrinder 2000) was used to grind the leaf tissues. The quality of DNA was not affected by long storage, and the quantity obtained per sample was adequate for about 1000 PCR reactions. Thus, this method allowed isolation of about 200 samples per day at a cost of $0.60 per sample and is a relatively high-throughput, low-cost extraction compared with conventional methods that use manual grinding and/or expensive kits.     

Effect of population density of compatible neighbours on inbreeding level within a Primula sieboldii population

We investigated the effect of population density of compatible neighbours on inbreeding level of Primula sieboldii, a heterostylous clonal herb. Pollinator availability, seed set, selfing rate, diversity of pollen donors, and fitness of progenies were compared between less and more isolated genets, which differed in the number of compatible opposite-morph genets within 20 m, the range at which most pollen flow occurred. Although pollinator availability did not differ between the two groups, seed set and diversity of pollen donors in more isolated genets were significantly lower than in less isolated genets. Additionally, the mean selfing rates of less and more isolated genets were 1.3 and 36.7%, respectively, and the mean leaf area of the self-fertilized seedlings was 70 to 40% smaller than that of outcrossed seedlings of the same mother genet. Due to this large inbreeding depression, it is unlikely that self-fertilized seedlings could successfully establish in natural habitats and hence the inbreeding level in the next generation around the more isolated genet would not increase rapidly. However, the possibility of mating between full-sibs would increase because the diversity of pollen donors was low and both pollen and seed dispersal were spatially restricted. Thus the inbreeding level of the next generations would gradually increase around the more isolated genets owing to biparental inbreeding. This study suggested that the population density of compatible neighbours has a critical impact on the future inbreeding level within P. sieboldii populations.     

Assessing wheat (Triticum aestivum L.) genetic diversity using quality traits, amplified fragment length polymorphisms, simple sequence repeats and proteome analysis

The genetic diversity among 10 Iranian bread wheat (Triticum aestivum) genotypes was analysed using 12 quality traits, 320 amplified fragment length polymorphisms (AFLP) polymorphic fragments, 491 simple sequence repeats (SSR) alleles and 294 proteome markers. The results revealed that the genotypes differed for quality traits, AFLP, SSR and proteome markers. The average genetic diversity based on quality traits (0.684 with a range of 0.266–0.997) was higher than AFLP (0.502 with a range of 0.328–0.717), SSR (0.503 with a range of 0.409–0.595) and proteome (0.464 with a range of 0.264–0.870) markers. Although there were apparent similarities between the groupings of particular genotypes, the overall correspondence between the distance matrices appeared to be rather low. In this study, the cluster analysis based on AFLP data showed the closest agreement with genotypes’ regions of origin or pedigree information. In addition to the genetic diversity assessment, specific proteins with known function were detected uniquely for the studied genotypes. Our results suggest that the classification based on quality traits and genotypic markers of these wheat genotypes will be useful for wheat breeders to plan crosses for positive traits.     

Isolation and characterization of microsatellite loci in Pimelodus microstoma (Siluriformes: Pimelodidae)

Ten microsatellite loci were developed and described for Pimelodus microstoma, identified in eggs collected from the lower Das Cinzas river. Nine loci were polymorphic, with 02–19 alleles per locus. The observed heterozygosity values were up to 0.898. Five species of the Pimelodidae family were also tested for amplification of loci, among which Pimelodus maculatus was the most successful in the transferability of primers. These loci have proven to be useful for studying the variability and structuring of populations of Pimelodus microstoma.     

基于转录组测序的茄子SSR标记开发

应用Trinity软件对茄子转录组测序数据组装,得到总长度为47919660 bp的45404条Unigenes,MISA从中检索到8316个SSR位点,发生频率为18.32%,平均5.63 kb一个位点。SSR位点中单碱基重复类型最多,为5372个,占到64.60%;其次为三碱基重复1628个,占到19.58%。三碱基重复中AAG/CTT是优势重复单元,占三碱基重复数的31.6%;二碱基重复中AG/CT是优势重复单元,占二碱基重复数的42.3%。利用Primer 3设计引物,共得到858对SSR引物,随机选取100对引物对17份茄子材料进行扩增,结果表明:有84对可以扩增出条带清晰的片段,有47对引物扩增片段为多态性片段。对47对多态性引物进行分析,多态性信息含量范围为0.10~0.64,平均多态性信息含量为0.32,UPGMA聚类分析可将17份材料分为3类。以上结果表明,基于茄子转录组测序开发的SSR标记可以为茄子的遗传多样性分析和遗传图谱构建提供更加丰富的标记来源。     

Genetic diversity and origin of weedy rice (Oryza sativa f. spontanea) populations found in North-eastern China revealed by simple sequence repeat (SSR) markers

BACKGROUND AND AIMS: Weedy rice (Oryza sativa f. spontanea) is one of the most notorious weeds occurring in rice-planting areas worldwide. The objectives of this study are to determine the genetic diversity and differentiation of weedy rice populations from Liaoning Province in North-eastern China and to explore the possible origin of these weedy populations by comparing their genetic relationships with rice varieties (O. sativa) and wild rice (O. rufipogon) from different sources. METHODS: Simple sequence repeat (SSR) markers were used to estimate the genetic diversity of 30 weedy rice populations from Liaoning, each containing about 30 individuals, selected rice varieties and wild O. rufipogon. Genetic differentiation and the relationships of weedy rice populations were analysed using cluster analysis (UPGMA) and principle component analysis (PCA). KEY RESULTS: The overall genetic diversity of weedy rice populations from Liaoning was relatively high (H(e) = 0.313, I = 0.572), with about 35 % of the genetic variation found among regions. The Liaoning weedy rice populations were closely related to rice varieties from Liaoning and japonica varieties from other regions but distantly related to indica rice varieties and wild O. rufipogon. CONCLUSIONS: Weedy rice populations from Liaoning are considerably variable genetically and most probably originated from Liaoning rice varieties by mutation and intervarietal hybrids. Recent changes in farming practices and cultivation methods along with less weed management may have promoted the re-emergence and divergence of weedy rice in North-eastern China.     

SSR and AFLP based genetic diversity of soybean germplasm differing in photoperiod sensitivity

Forty-four soybean genotypes with different photoperiod response were selected after screening of 1000 soybean accessions under artificial condition and were profiled using 40 SSR and 5 AFLP primer pairs. The average polymorphism information content (PIC) for SSR and AFLP marker systems was 0.507 and 0.120, respectively. Clustering of genotypes was done using UPGMA method for SSR and AFLP and correlation was 0.337 and 0.504, respectively. Mantel's correlation coefficients between Jaccard's similarity coefficient and the cophenetic values were fairly high in both the marker systems (SSR = 0.924; AFLP = 0.958) indicating very good fit for the clustering pattern. UPGMA based cluster analysis classified soybean genotypes into four major groups with fairly moderate bootstrap support. These major clusters corresponded with the photoperiod response and place of origin. The results indicate that the photoperiod insensitive genotypes, 11/2/1939 (EC 325097) and MACS 330 would be better choice for broadening the genetic base of soybean for this trait.     

Simple sequence repeat analyses of interspecific hybrids and MAALs of <Emphasis Type="Italic">Oryza </Emphasis><Emphasis Type="Italic">officinalis</Emphasis> and <Emphasis Type="Italic">Oryza sativa</Emphasis>

Wild rice is a valuable resource for the genetic improvement of cultivated rice (Oryza sativa L., AA genome). Molecular markers are important tools for monitoring gene introgression from wild rice into cultivated rice. In this study, Simple sequence repeat (SSR) markers were used to analyze interspecific hybrids of O. sativa-O. officinalis (CC genome), the backcrossing progenies and the parent plants. Results showed that most of the SSR primers (335 out of 396, 84.6%) developed in cultivated rice successfully amplified products from DNA samples of wild rice O. officinalis. The polymorphism ratio of SSR bands between O. sativa and O. officinalis was as high as 93.9%, indicating differences between the two species with respect to SSRs. When the SSR markers were applied in the interspecific hybrids, only a portion of SSR primers amplified O. officinalis-specific bands in the F(1) hybrid (52.5%), BC(1) (52.5%), and MAALs (37.0%); a number of the bands disappeared. Of the 124 SSR loci that detected officinalis-specific bands in MAAL plants, 96 (77.4%) showed synteny between the A and C-genomes, and 20 (16.1%) showed duplication in the C-genome. Sequencing analysis revealed that indels, substitution and duplication contribute to the diversity of SSR loci between the genomes of O. sativa and O. officinalis.