Genome assembly of primitive cultivated potato Solanum stenotomum provides insights into potato evolution.

Genetic diversity is the raw material for germplasm enhancement. Landraces and wild species relatives of potato, which contain a rich gene pool of valuable agronomic traits, can provide insights into the genetic diversity behind the adaptability of the common potato. The diploid plant, Solanum stenotomum (Sst), is believed to have an ancestral relationship with modern potato cultivars and be a potential source of resistance against disease. Sequencing of the Sst genome generated an assembly of 852.85 Mb (N50 scaffold size, 3.7 Mb). Pseudomolecule construction anchored 788.75 Mb of the assembly onto 12 pseudochromosomes, with an anchor rate of 92.4%. Genome annotation yielded 41,914 high-confidence protein-coding gene models and comparative analyses with closely related Solanaceae species identified 358 Sst-specific gene families, 885 gene families with expansion along the Sst lineage, and 149 genes experiencing accelerated rates of protein sequence evolution in Sst, the functions of which were mainly associated with defense responses, particularly against bacterial and fungal infection. Insights into the Sst genome and the genomic variation of cultivated potato taxa are valuable in elaborating the impact of potato evolution in early landrace diploid and facilitate modern potato breeding.

Optimal assembly strategies of transcriptome related to ploidies of eukaryotic organisms.

BACKGROUND: Several de novo transcriptome assemblers have been developed recently to assemble the short reads generated from the next-generation sequencing platforms and different strategies were employed for assembling transcriptomes of various eukaryotes without genome sequences. Though there are some comparisons among these de novo assembly tools for assembling transcriptomes of different eukaryotic organisms, there is no report about the relationship between assembly strategies and ploidies of the organisms. RESULTS: When we de novo assembled transcriptomes of sweet potato (hexaploid), Trametes gallica (a diploid fungus), Oryza meyeriana (a diploid wild rice), five assemblers, including Edena, Oases, Soaptrans, IDBA-tran and Trinity, were used in different strategies (Single-Assembler Single-Parameter, SASP; Single-Assembler Multiple-Parameters, SAMP; Combined De novo Transcriptome Assembly, CDTA, that is multiple assembler multiple parameter). It was found that CDTA strategy has the best performance compared with other two strategies for assembling transcriptome of the hexaploid sweet potato, whereas SAMP strategy with assembler Oases is better than other strategies for assembling transcriptomes of diploid fungus and the wild rice transcriptomes. CONCLUSION: Based on the results from ours and others, it is suggested that CDTA strategy is better used for transcriptome assembly of polyploidy organisms and SAMP strategy of Oases is outperformed for those diploid organisms without genome sequences.

Modulated expression of genes associated with NO signal transduction contributes to the cholesterol-lowering effect of electro-acupuncture.

Electro-acupuncture (EA) at Fenglong acupoint (ST40) can lower the levels of serum cholesterol and triacylglycerols. To study the hepatic genes responsible for the cholesterol-lowering effect of EA, suppression subtractive hybridization combined with the switch mechanism at the 5'-end of RNA template cDNA synthesis and long-distance PCR were employed using hepatic tissues from hypercholesterolemia and EA-treated mice. 68 % of the identified genes are involved in metabolism, immune response, and signal transduction pathways. Real-time PCR and western blot indicate that EA at ST40 induces the expression of nNOS and Mt1, two genes involved in NO signal transduction. EA treatment for hypercholesterolemia thus involves the modulation of several biological pathways and provides a physiological link between NO signal transduction and the cholesterol-lowering effect of EA.

Tea catechins induce the conversion of preformed lysozyme amyloid fibrils to amorphous aggregates.

Natural polyphenols are major constituents of plant foods and herbs. Numerous studies have demonstrated that natural polyphenols inhibited amyloid formation and destabilized the preformed amyloid fibrils. However, the molecular mechanism for the antiamyloidogenesis of polyphenols is still unclear and remains to be further explored. In the present study, the preformed lysozyme fibrils were used as an in vitro model to study the disruptive effects of tea catechins on amyloid fibrils. Results showed that tea catechins induced the conversion of lysozyme fibrils to amorphous aggregates and inhibited fibril-induced hemolysis. Hydroquinone also showed disruptive effect on the fibrils, whereas phenol and two typical antioxidants, ascorbic acid and alpha-tocopherol, did not affect the fibrillar structure, suggesting that polyphenolic structure is essential for fibril deposition. Correlation analyses indicate that the fibril-depositing effects were related to both the antioxidative potency and hydrophobicity of tea catechins. These findings provide new evidence for comprehensive understanding of the interaction between natural polyphenols and amyloid fibrils.

Identification and characterization of differentially expressed genes involved in pharmacological activities of roots of Panax notoginseng during plant growth.

Panax notoginseng is a highly valued Chinese medicinal herb. To understand the molecular mechanism of the much higher pharmacological activities of roots of 3-year-old plants over 1-year-old ones,two cDNA libraries were constructed using the suppression subtractive hybridization (SSH) method. Positive cDNA clones from each of the two libraries were randomly selected for dot-blotting analysis. A total of 110 genes were highly expressed in 3-year-old roots and 80 genes in 1-year-old roots. Of these, 87 cDNA fragments were sequenced, assembled, and compared with sequences in GenBank, and 81 individual cDNAs were identified. These cDNAs were the first expressed sequence tags of P. notoginseng in GenBank. The result of reverse transcription PCR analysis of six genes was consistent with that of the dot-blot analysis. The global gene expression profile showed that there were significant differences between 1- and 3-year-old roots of P. notoginseng plants. Some important structural and regulatory genes which may be involved in isoprenoid biosynthesis were found to be over-expressed in 3-year-old roots, such as genes encoding 1-hydroxy-2-methyl-2-(E)-butenyl-4-diphosphat-synthase (IspG-protein), multi-copper oxidase type I family protein, NADH flavin oxidoreductase, lipase and aconitase.

Molecular cloning and characterization of the gene encoding squalene epoxidase in Panax notoginseng.

Squalene epoxidase (SE) is one of the rate-limiting enzymes in the triterpene saponins biosynthetic pathway. Panax notoginseng, one of the famous medicinal plants in China, produces bioactive triterpene saponins. Here we report the P. notoginseng SE, which was cloned from the root of P. notoginseng by PCR. The nucleotide sequence of the ORF (GenBank accession no. DQ386734) contains 1611 nucleotides and encodes 537 amino acid residues with molecular weight of 59.14 kDa and pI of 8.81. The gene has 98% identity with P. ginseng but different identities with other SE families. P. notoginseng SE has a FAD function domain, NAD(P)-binding Rossmann-fold domains, hydrophobicity and 4 transmembrane helices. This SE may be a microsomal membrane-associated enzyme. Real time quantitative PCR shows that the cDNA has different expression pattern and is highly expressed in root, especially in 3-year-old root.

Electroacupuncture alters the expression of genes associated with lipid metabolism and immune reaction in liver of hypercholesterolemia mice.

Acupuncture or electroacupuncture (EA) is effective in treating various metabolism disorders. Previously we found that EA at the acupoint, Fenglong (ST40), had the cholesterol-lowering effect and regulated genes expression in liver of hypercholesterolemia mice (M Li and YZ Zhang, Int J Mol Med 2007, 19: 617-629). To explain gene expression associated with EA, suppression subtractive hybridization (SSH), combined with targeted display (TD), was used and 26 up-regulated and 24 down-regulated genes with known functions were identified in hypercholesterolemia mice liver, some of which are involved in key reactions of lipid metabolism and immune reaction. Promoting lipid metabolism and suppressing inflammation via modulating mRNA expression may be the mechanism of EA inducing modulation of cholesterol concentrations.

Modulation of gene expression in cholesterol-lowering effect of electroacupuncture at Fenglong acupoint (ST40): a cDNA microarray study.

The aim of this study was to demonstrate the cholesterol-lowering effect of electroacupuncture (EA) at the acupoint of Fenglong (ST40) in mice and to investigate its molecular mechanism by using genome-wide gene expression profile analysis. Mice with hypercholesterolemia induced by a high-cholesterol diet were randomly divided into EA at ST40 group (EG), EA at non-acupoint group (ENG), and simvastatin group (DG). A lipid profile of both the plasma and liver indicated that EA at ST40 had the same hypocholesterolemic effect as that of simvastatin, while EA at non-acupoint failed to produce the same effect. The global gene expression profile showed that EA at ST40 not only regulated the expression of genes which were directly involved in the cholesterol metabolism in the liver, but also significantly affected the expression of genes involved in signal transduction, transcription regulation, cell cycle, cell adhesion, immunity and stress. The gene expression pattern was further verified by real-time RT-PCR. The mechanism by which EA at ST40 regulated liver cholesterol metabolism is discussed. We conclude that the hypocholesterolemic effect is specific to EA at ST40 and not due to general electrical stimulation of muscles. The comprehensive gene expression profile analysis appears particularly useful in the search for EA-induced changes in cholesterol regulation.

Molecular cloning and expression of yak (Bos grunniens) lactoferrin cDNA in Pichia pastoris.

cDNA encoding lactoferrin from yak was isolated by RT-PCR and then sequenced. The cloned cDNA (2127 bp) encodes a 709 amino acid precursor molecule of yak lactoferrin with a signal peptide of 19 amino acids. The yak lactoferrin cDNA was expressed in Pichia pastoris. The recombinant protein, purified by Ni-NTA affinity column, had a molecular weight of 76 kDa and reacted with an antibody raised against native bovine lactoferrin. The iron-binding behavior and antimicrobial activity of the purified protein indicated that it was correctly folded and functional.

Isolation of genes involved in the preventive effect of electroacupuncture at Fenglong acupoint (ST40) on hypercholesterolemia mice by suppression subtractive hybridization (SSH) combined with negative subtraction chain (NSC) technology.

We have shown that electroacupuncture (EA) at Fenglong acupoint (ST40) has the cholesterol-lowering effect in hypercholesterolemia mice. The present study was designed to study preventive effect of EA at ST40 on hypercholesterolemia. C57BL/6j mice were randomly divided into normal group (NG), hypercholesterolemia group (HG) and EA prevention group (EPG). NG were fed chow, HG a hypercholesterolemic diet (HD), and EPG the same HD and received EA treatment simultaneously. Lipid profile of both the plasma and liver indicated that EA at ST40 had preventive effect on hypercholesterolemia. Compared with corresponding values in the HG mice, the levels of the hepatic total cholesterol and total triglyceride in the EPG mice lowered 45% and 23% respectively, and the levels of plasma total-, LDL-, and HDL-cholesterol in the EPG mice lowered 39%, 37% and 39% respectively. Eleven genes whose expressions were up-regulated in EPG mice compared with HG were isolated using suppression subtractive hybridization (SSH) combined with negative subtraction chain (NSC) technology, and then confirmed by dot-blot assay. Except two genes whose functions were still unknown, the others were mainly involved in cholesterol metabolism, lipid metabolism, glucose metabolism and immune response. The potential molecular mechanism of preventive effect was discussed.