Association of T2/S-RNase With Self-Incompatibility of Japanese Citrus Accessions Examined by Transcriptomic, Phylogenetic, and Genetic Approaches

Several citrus varieties show gametophytic self-incompatibility (GSI), which can contribute to seedless fruit production in several cultivars. This study investigated the genes regulating this trait through RNA-seq performed using styles collected from the flowers of Japanese citrus cultivars ‘Hyuganatsu,' ‘Tosabuntan,' ‘Hassaku,' ‘Banpeiyu,' and ‘Sweet Spring'. We screened the transcripts of putative T2 RNases, i.e., the protein family including all S-RNases from S-RNase-based GSI plants, and constructed a phylogenetic tree using the screened T2 RNases and S-RNases retrieved from citrus genome databases and a public database. Three major clusters (class I–III) were formed, among which, the class III cluster contained family specific subclusters formed by S-RNase and a citrus-specific cluster monophyletic to the S-RNase clusters. From the citrus class III cluster, six transcripts were consistent with the S haplotypes previously determined in Japanese citrus accessions, sharing characteristics such as isoelectric point, extracellular localization, molecular weight, intron number and position, and tissue-specific expression with S-RNases. One T2 RNase gene in self-incompatible Hyuganatsu was significantly down-regulated in the styles of a self-compatible mutant of Hyuganatsu in RNA-seq and qPCR analyses. In addition, the inheritance pattern of some T2 RNase genes was consistent with the pattern of the S haplotype in the progeny population of Hyuganatsu and Tosabuntan. As all results supported citrus self-incompatibility being based on S-RNase, we believe that six T2 RNase genes were S-RNases. The homology comparison between the six T2 RNases and S-RNases recently reported in Chinese citrus revealed that three out of six T2 RNases were identical to S-RNases from Chinese citrus. Thus, the other three T2 RNases were finally concluded to be novel citrus S-RNases involved in self-incompatibility.

Microarray Analysis Reveals a Potential Role of lncRNA Expression in remote ischemic preconditioning in Myocardial ischemia-reperfusion injury

Background Avoiding or reducing cardiopulmonary bypass-related injuries of important organs is the eternal topic in the field of cardiovascular surgery. Remote ischemic preconditioning (RIPC) is a promising strategy whose clinical application seems more realistic and extensive compared with other conservative or surgical strategies. However, considering its complex process which involves the expression of a variety of molecules and even genes and little is known about its underlying mechanism after years of researches, new train of thought and method must be developed to explore its potential in clinical application. Long noncoding RNA (LncRNA) is a kind of RNA that has been proven be exert efficacy in the occurrence and development of cardiovascular disease. The differential expression of LncRNAs and their biological effects during RIPC have not been reported. Methods We first verified the protective effect of RIPC on myocardial ischemia-reperfusionby western blot, ELISA and hematoxylin-eosin(HE) staining. Mouse and human LncRNA arrays were used to investigate the expression signatures of LncRNAs and mRNAs in myocardial tissue after RIPC. Homology comparison was used to screen homologous genes in total differentially expressed LncRNAs. Competing endogenous RNA(ceRNA) mechanism analysis helped us find the matching relationship among homologous LncRNA, mRNA and microRNA. Results 554 differentially expresssed mouse LncRNAs(281 up-regulated / 273 down-regulated) and 1392 differentially expresssed human LncRNAs(635 up-regulated / 757 down-regulated) were selected from LncRNA microarrays. Quantitative real-time polymerase chain reaction(qRT-PCR) was used to verify these selected LncRNAs, after homology comparison and ceRNA mechanism analysis, we got a pair of homologous LncRNAs(ENST00000574727 & ENSMUST00000123752), predicted corresponding matching relationship and potential competitive binding sites. Conclusions In this study, we identify a number of differentially expressed LncRNAs, which may be closely related to the regulation of inflammation and cell proliferation. In addition, we take several pairs of homologous LncRNAs as the next research object. By continuously exploring the function and mechanism of these LncRNAs, we hope to unveil the mystery of RIPC and discover a new protective mechanism of cardiovascular ischemia-reperfusion disease.

A Computational Methodology to Overcome the Challenges Associated With the Search for Specific Enzyme Targets to Develop Drugs Against Leishmania major

We present an approach for detecting enzymes that are specific of Leishmania major compared with Homo sapiens and provide targets that may assist research in drug development. This approach is based on traditional techniques of sequence homology comparison by similarity search and Markov modeling; it integrates the characterization of enzymatic functionality, secondary and tertiary protein structures, protein domain architecture, and metabolic environment. From 67 enzymes represented by 42 enzymatic activities classified by AnEnPi (Analogous Enzymes Pipeline) as specific for L major compared with H sapiens, only 40 (23 Enzyme Commission [EC] numbers) could actually be considered as strictly specific of L major and 27 enzymes (19 EC numbers) were disregarded for having ambiguous homologies or analogies with H sapiens. Among the 40 strictly specific enzymes, we identified sterol 24-C-methyltransferase, pyruvate phosphate dikinase, trypanothione synthetase, and RNA-editing ligase as 4 essential enzymes for L major that may serve as targets for drug development.

Research of the Unknown Cause of Death of Caesio diagramma

To find out the etiology and prevention methods of the death ofCaesio diagramma. Isolate the pathogen from the fester surface, liver, kidney , intestinal and other organizations of the diseasedCaesio diagramma; After the purification of the predominant strains, the phylogenetic analysis of genetic relationships was done by determining 16s rDNA gene and Homology comparison was done with the BLAST sequence detected and registered in GenBank sequence ; In the end, phylogenetic trees were constructed and the drug sensitivity experiment and pathological observation were taken.Aeromonas hydrophilais the cause of the death ofCaesio diagramma, it is highly sensitive to chloramphenicol, florfenicol, cefotaxime, gentamicin, amikacin; The pathological results indicate that the wax buildups causes increasing energe consumption ofCaesio diagramma,which decreased the resistance ofCaesio diagrammaand even leads to secondary bacterial infection. Pathogen infected the liver and spleen tissues which causes the disease and death ofCaesio diagramma.Aeromonas hydrophilacauses the death ofCaesio diagramma, and the results have an important guiding significance for the healthy breeding ofCaesio diagrammaand other ornamental fish

Metagenomic Analysis of the Viral Communities in Fermented Foods

ABSTRACTViruses are recognized as the most abundant biological components on Earth, and they regulate the structure of microbial communities in many environments. In soil and marine environments, microorganism-infecting phages are the most common type of virus. Although several types of bacteriophage have been isolated from fermented foods, little is known about the overall viral assemblages (viromes) of these environments. In this study, metagenomic analyses were performed on the uncultivated viral communities from three fermented foods, fermented shrimp, kimchi, and sauerkraut. Using a high-throughput pyrosequencing technique, a total of 81,831, 70,591 and 69,464 viral sequences were obtained from fermented shrimp, kimchi and sauerkraut, respectively. Moreover, 37 to 50% of these sequences showed no significant hit against sequences in public databases. There were some discrepancies between the prediction of bacteriophages hosts via homology comparison and bacterial distribution, as determined from 16S rRNA gene sequencing. These discrepancies likely reflect the fact that the viral genomes of fermented foods are poorly represented in public databases. Double-stranded DNA viral communities were amplified from fermented foods by using a linker-amplified shotgun library. These communities were dominated by bacteriophages belonging to the viral orderCaudovirales(i.e.,Myoviridae,Podoviridae, andSiphoviridae). This study indicates that fermented foods contain less complex viral communities than many other environmental habitats, such as seawater, human feces, marine sediment, and soil.

ESTs from Seeds to Assist the Selective Breeding of Jatropha curcas L. for Oil and Active Compounds

We report here on the characterization of a cDNA library from seeds of Jatropha curcas L. at three stages of fruit maturation before yellowing. We sequenced a total of 2200 clones and obtained a set of 931 non-redundant sequences (unigenes) after trimming and quality control, ie, 140 contigs and 791 singlets with PHRED quality >10. We found low levels of sequence redundancy and extensive metabolic coverage by homology comparison to GO. After comparison of 5841 non-redundant ESTs from a total of 13193 reads from GenBank with KEGG, we identified tags with nucleotide variations among J. curcas accessions for genes of fatty acid, terpene, alkaloid, quinone and hormone pathways of biosynthesis. More specifically, the expression level of four genes (palmitoyl-acyl carrier protein thioesterase, 3-ketoacyl-CoA thiolase B, lysophosphatidic acid acyltransferase and geranyl pyrophosphate synthase) measured by real-time PCR proved to be significantly different between leaves and fruits. Since the nucleotide polymorphism of these tags is associated to higher level of gene expression in fruits compared to leaves, we propose this approach to speed up the search for quantitative traits in selective breeding of J. curcas. We also discuss its potential utility for the selective breeding of economically important traits in J. curcas.

(286) Identification of Genes in Tomato Roots Induced by Aluminum Stress using Fluorescent cDNA Differential Display

The goal of this study was isolate genes that are regulated by Al treatments of tomato roots growing in vitro. For Al treatment, germinating tomato seeds were plated on a MS agar medium supplemented with 0, 350, and 1200 μM AlCl3 for 30 days. Total RNA was extracted from root tissues and separated on denature formamide gel to check their quantity and quality. Equal amount of total RNA from treatment and control was treated with DNAse I (Genhunter, TN) to remove genomic DNA contamination. cDNA was obtained by reverse transcription using all the regents in RNA Image Kit (Genhunter, TN). The cDNA was amplified using the fluorescently labeled anchor primers (Oligo dT-A, C, G) and 16 random primers. Amplification products were separated by electrophoresis in 6% nondenaturing polyacrylamide gels and DNA bands were observed by scanning the gel on a FMBIOIII scanner. After comparing the band profile on the gel image, fragments of gene that showed changes in intensity compared to control (0 μM AlCl3) were isolated from the gel manually. These bands were re-amplified with the same pair of primers as the original amplification and cloned onto PCR-Trap cloning vector (Genhunter, TN). After DNA sequence analysis and homology comparison with NCBI database, we have identified clone # C01HBa0256E08 on L. esculentum chromosome 01, clone # C10HBa0111D09 on chromosome 10 and clone # LE_HBa-31H5 on chromosome 4.