High Levels Of Polysaccharides Research Articles

A Comparative Study on Macronutrients Content and Antioxidant Activity of Four Wild and Cultivated Seaweeds from the Moroccan Atlantic Coast

ABSTRACT: Extensive harvest of seaweeds in Morocco has caused overexploitation of these resources. Four Moroccan common seaweed species, wild and cultivated, were studied to assess the cultivated species' potential in comparison to their wild counterparts. The macronutrient composition, total phenolic compounds (TPC), as well as potential antioxidant activity (DPPH and FRAP), were established. Seaweeds contained a wide range of proteins (5–16% of DW), lipids (1–3% of DW), carbohydrates (20–59% of DW), and total polyphenols (3-6 mg EAG/g of DW). Red seaweeds contained high levels of polysaccharides (47–59% of DW), mainly the cultivated Gigartina pistillata, and important amounts of proteins, especially Gelidium sesquipedale (14–15% of DW), whereas the cultivated type was richer in proteins. Gracilaria gracilis and wild Gelidium sesquipedale are characterized by notable antioxidant activity, particularly the cultivated Gracilaria gracilis (TPC: 6.807 mg GAE/g of DW; low values of IC50 of DPPH: 53.863μg/mL and FRAP: 67.033μg/mL). Besides, the green seaweed Ulva lactuca is poor in carbohydrates but contains essential amounts of proteins and phenolics. This work highlights the significant potential of cultivating Moroccan seaweeds through algaculture to achieve more sustainable production with enhanced nutrient and bioactive molecule content.

Sulfur bacteria-reinforced microbial electrochemical denitrification

Two limiting factors of microbial electrochemical denitrification (MED) are the abundance and efficiency of the functional microorganisms. To supply these microorganisms, MED systems are inoculated with denitrifying sludge, but such method has much room for improvement. This study compared MED inoculated with autotrophic denitrifying inoculum (ADI) versus with heterotrophic denitrifying inoculum (HDI). ADI exhibited electroactivity for 50% less of timethan HDI. The denitrification efficiency of the ADI biocathode was42% higherthan that of the HDI biocathode. The HDI biocathode had high levels of polysaccharides while the ADI biocathode was rich in proteins, suggesting that two biocathodes may achieveMED but via differentpathways. Microbial communities of two biocathodes indicated MED of HDI biocathode may rely on interspecies electron transfer, whereas sulfur bacteria of ADI biocathode take electrons directly from the cathode to achieve MED. Utilizing autotrophic sulfur-oxidizing denitrifiers, this study offers a strategy for enhancing MED.

Comparison and improvement of RNA extraction methods in Sargassum (Phaeophyta).

Sargassum (Sargassaceae) is widely distributed globally and plays an important role in regulating climate change, but the landscape of genomes and transcripts is less known. High-quality nucleic acids are the basis for molecular biology experiments such as high-throughput sequencing. Although extensive studies have documented methods of RNA extraction, these methods are not very applicable to Sargassum, which contains high levels of polysaccharides and polyphenols. To find a suitable method to improve the quality of RNA extracted, we compared and modified several popular RNA extraction methods and screened one practical method with three specific Sargassum spp. The results showed that three CTAB methods (denoted as Methods 1, 2, and 3) and the RNAprep Pure Plant Kit (denoted as Method 4) could, with slight modifications, effectively isolate RNA from Sargassum species, except for Method 4 used with S. fusiforme. By performing further screening, we determined Method 4 was the best choice for S. hemiphyllum and S. henslowianum, as revealed by RNA yields, RNA Integrity Number (RIN), extraction time, and unigene mapped ratio. For S. fusiforme, Methods 1, 2, and 3 showed no obvious differences among the yields, quality, or time to perform. In addition, one other method was tested, but we found the quality of the RNA extracted by TRIzol reagent methods (denoted as Method 5) performed the worst when compared with the above four methods. Therefore, our study provides four suitable methods for RNA extraction in Sargassum and is essential for future genetic exploration of Sargassum.

High-quality DNA isolation protocol for detection of Khapra beetle (Dermestidae: Trogoderma granarium Everts, 1898) in standard wheat germ trap.

Khapra beetle (Dermestidae: Trogoderma granarium Everts, 1898) is an internationally significant pest of grain crops and stored grain products. Wheat germ traps, routinely used in surveillance sampling of Khapra beetle provide feed-substrates used by the pest throughout its life cycle. However, Khapra beetle larvae, eggs and other traces of the pest, such as larval frass and exuviae, in wheat germ traps are difficult to sort and taxonomically identify. Additionally, high levels of polysaccharides in wheat germ can inhibit PCR based molecular detection of this pest captured in the traps. We have developed a sensitive and low-cost protocol for extracting trace levels of Khapra beetle DNA from an entire wheat germ trap. Overnight digestion of entire trap contents in 6 mL of ATL buffer, followed by a 40min lysis step was optimal for DNA extraction. Paired with reported qPCR assays, this protocol allows the detection of a few hairs of T. granarium in a typical 2-gram wheat germ trap. This DNA extraction protocol makes it possible to perform a more rapid identification of the pest following wheat germ sample collection. The protocol has potential to improve international efforts for Khapra beetle surveillance.

Genome sequencing and analysis of Xylaria sp. L1 provides novel insights into its utilization in IDF decomposition in okara

This study investigated the genome ofXylaria sp. L1 and its application in assisting the degradation of okara, which is an agricultural waste. The fungus process showed that there was a 50.31% cellulose loss and 56.25% hemicellulose loss, respectively. In merely 7 days, compared with untreated control, the content of insoluble dietary fiber (IDF) in the substrate was significantly decreased by 62% whereas that of soluble dietary fiber (SDF) significantly increased by 413%. Importantly, the activity of cellulase showed a continuous increase trend and it reached the maximum activity at 11.37U/g on the 7th day. The pretreated biomass showed much higher levels of polysaccharides, especially for glucuronic acid (63.63%), than the untreated controls after 7 days. Additionally, Xylaria sp. L1 sporocarps had higher quantities of macronutrients, especially for Zn. Furthermore, we conducted the first annotation of Xylaria sp. L1 genome. Illumina and nanopore sequencing revealed that the genome size of Xylaria sp. L1 was 36.61 Mb. The high activity of cellulase was in accordance with the annotation the Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), Eukaryotic Orthologous Groups (KOG), and carbohydrate-active enzymes (CAZy) databases, which are related to cellulose and hemicellulose degradation. GHs were the most abundant enzymes and 203 genes were annotated to GHs. Cellulose and hemicellulose decomposition was likely performed by an expressed GH43 and GH5 protein families containing their binding domains and genes A0779, A0819, A7717 played an important role in pentose and glucuronate interconversions. Therefore, the higher IDF decomposition by Xylaria sp. L1 makes it a prospective feedstock for okara degradation.

Oral Wound Healing Potential of Polygoni Cuspidati Rhizoma et Radix Decoction-In Vitro Study.

Polygoni Cuspidati Rhizoma et Radix (syn. rhizomes of Reynoutria japonica Houtt.) is a pharmacopoeial raw material in Europe and China. In traditional medicine, one of the applications for Reynoutria japonica rhizomes is wound healing. In a recent in vitro study, we demonstrated that ethanol and acetone extracts from this herbal drug have the potential to heal oral gum wounds. However, considering that a majority of herbal medicines have been traditionally administered as water decoctions, in the present study, a decoction of Reynoutria japonica rhizomes was prepared and detailed tests to determine its in vitro gingival wound healing activity were conducted. We used the primary human gingival fibroblasts (HGF) incubated with a decoction to determine cell viability (MTT assay), cell proliferation (the confocal laser scanning microscope-CLSM), and cell migration (wound healing assay). Moreover, the collagen type III expression was examined using immunocytochemical staining. The studied decoction was qualitatively and quantitatively characterized using the validated HPLC/DAD/ESI-HR-QTOF-MS method. The Folin-Ciocalteu assay was used to determine the total phenols and tannins content. Additionally, HPLC-RI analysis of decoction and the previously obtained ethanol and acetone extracts was used to determine the composition of saccharides. Low concentration (from 50 to 1000 µg/mL) of decoction after 24 h incubation caused a significant increase in HGF cell viability. No cytotoxic effect was observed at any tested concentration (up to 2000 µg/mL). The lowest active concentration of decoction (50 µg/mL) was selected for further experiments. It significantly stimulated human gingival fibroblasts to proliferate, migrate, and increase the synthesis of collagen III. Phytochemical analysis showed significantly fewer polyphenols in the decoction than in the ethanol and acetone extracts tested earlier. In contrast, high levels of polysaccharides were observed. In our opinion, they may have a significant effect on the oral wound healing parameters analyzed in vitro. The results obtained encourage the use of this raw material in its traditional, safe form-decoction.

Genomic DNA Extraction: The Critical Stage in Plant Molecular Analysis Activities

<p>DNA extraction is a crucial stage in plant molecular analysis activities that determine the success of further downstream analysis stages. Molecular analysis activities, including Polymerase Chain Reaction (PCR), restriction digestion, DNA hybridisation, genomic library construction, and DNA sequencing, mostly require DNA with high quality and enough quantity. Some plants contain high levels of polysaccharides, proteins, and secondary metabolites, such as polyphenols, tannins, and alkaloids, which can affect the purity of the isolated DNA. The presence of these contaminants can inhibit the activity of several enzymes, such as DNA polymerase and restriction enzymes, so that DNA could not be amplified or digested. Various DNA extraction methods have been developed, ranging from manual method, manual commercial kits, and robotic techniques. Each method has its advantages and disadvantages since the selection of the right method needs to be determined carefully in order to obtain the best results. Some modifications at certain stages in DNA extraction process need to be conducted to minimize the number of DNA contaminants. Several rapid DNA extraction methods nowadays have also been developed to save times, labors, and costs. In addition, the rapid DNA extraction method could also reduce the use of some hazardous chemical compounds, so that the method is not only giving economic benefits, but also good for human health.</p>

Polysaccharides from Callerya speciosa alleviate metabolic disorders and gut microbiota dysbiosis in diet-induced obese C57BL/6 mice.

Callerya speciosa ("Niu Dali" in Chinese) is a well-known edible plant in Southeast China. C. speciosa roots contain a high level of polysaccharides, which have been reported to show multiple health-promoting effects. In the current study, the anti-obesity effects of a crude extract of C. speciosa polysaccharides (NP) and its underlying mechanisms of action are investigated. C57BL/6 mice were divided into three groups and fed either a standard diet or a high-fat diet (HFD). The HFD + NP group mice received oral administration of NP (100 mg per kg per day) every other day for 10 weeks. NP supplementation alleviated HFD-induced diabetic biomarkers including body weight gain, hyperlipidemia, liver steatosis, and adipocyte hypertrophy. Western blot and RT-PCR analyses revealed that NP inhibited hepatic de novo lipogenesis and adipogenesis (i.e. decreased expression of Srebp1c, Fas, Cebpα, and Pparγ), stimulated adipocyte lipolysis (enhanced mRNA expression of Hsl and Mgl), and attenuated HFD-induced hepatic inflammation (decreased expression of TNF-α and NF-κB p65). Furthermore, 16S rDNA and GC-MS analyses showed that NP supplementation restored the Firmicutes/Bacteroidetes proportion, elevated colon-derived SCFAs, especially acetic acid content, and increased the relative abundance of genera associated with SCFA production in HFD-fed mice. Findings from this study suggest that NP alleviated HFD-induced obesity in a mouse model, which was possibly due to its ameliorative effects on diet-induced gut dysbiosis. Polysaccharides from C. speciosa are promising prebiotics and they may be further developed as functional foods for the management of obesity.

A native chromatin immunoprecipitation (ChIP) protocol for studying histone modifications in strawberry fruits

BackgroundCovalent modifications of histones and histone variants have great influence on chromatin structure, which is involved in the transcriptional regulation of gene expression. Chromatin immunoprecipitation (ChIP) is a powerful tool for studying in vivo DNA-histone interactions. Strawberry is a model for Rosaceae and non-climacteric fruits, in which histone modifications have been implicated to affect fruit development and ripening. However, a validated ChIP method has not been reported in strawberry, probably due to its high levels of polysaccharides which affect the quality of prepared chromatin and the efficiency of immunoprecipitation.ResultsWe describe a native chromatin immunoprecipitation (N-ChIP) protocol suitable for strawberry by optimizing the parameters for nuclei isolation, chromatin extraction, DNA fragmentation and validation analysis using quantitative real-time PCR (qRT-PCR). The qRT-PCR results show that both the active mark H3K36me3 and the silent mark H3K9me2 are efficiently immunoprecipitated for the enriched regions. Compared to X-ChIP (cross-linked chromatin followed by immunoprecipitation), our optimized N-ChIP procedure has a higher signal-to-noise ratio and a lower background for both the active and the silent histone modifications. Furthermore, high-throughput sequencing following N-ChIP demonstrates that nearly 90% of the enriched H3K9/K14ac peaks are overlapped between biological replicates, indicating its remarkable consistency and reproducibility.ConclusionsAn N-ChIP method suitable for the fleshy fruit tissues of woodland strawberry Fragaria vesca is described in this study. The efficiency and reproducibility of our optimized N-ChIP protocol are validated by both qRT-PCR and high-throughput sequencing. We conclude that N-ChIP is a more suitable method for strawberry fruit tissues relative to X-ChIP, which could be combined with high-throughput sequencing to investigate the impact of histone modifications in strawberry and potentially in other fruits with high content of polysaccharides.

Commercial Saccharomyces cerevisiae Yeast Strains Significantly Impact Shiraz Tannin and Polysaccharide Composition with Implications for Wine Colour and Astringency

To gain knowledge on the role of Saccharomyces cerevisiae yeast strains (and their hybrids) on wine sensory properties, 10 commercially available yeast strains were selected on the basis of their widespread usage and/or novel properties and used to produce Shiraz wines. Significant differences were evident post-alcoholic fermentation and after 24 months of ageing with regards to the number of wine compositional variables, in particular the concentration of tannin and polysaccharide. Strain L2323 is known for its pectinolytic activity and yielded the highest concentration of both yeast- and grape-derived polysaccharides. Wines made with the mannoprotein-producing strain Uvaferm HPS (high levels of polysaccharides) did not have elevated concentrations of yeast-derived polysaccharides, despite this observation being made for corresponding model fermentations, suggesting that mannoprotein production or retention might be limited by the wine matrix. Wine tannin concentration showed a high level of variability between strains, with L2323 having the highest, and AWRI1503 the lowest concentration. Sensory analysis of the wines after 24 months ageing revealed significant differences between the yeast strains, but only the attributes opacity (visual colour) and astringency could be predicted by partial least squares regression using the wine compositional data. Notably, the astringency attribute was associated with higher concentrations of both tannin and polysaccharide, contrary to reports in the literature which suggested that polysaccharide exerts a moderating effect on astringency. The results confirm previous reports demonstrating that the choice of yeast strain represents an opportunity to shape wine style outcomes.

A rapid and efficient SDS-based RNA isolation protocol from different tissues of coffee.

Isolation of high-quality RNA from coffee is challenging because of high level of polysaccharides, polyphenols and other secondary metabolites. In the present study, a rapid and efficient RNA extraction protocol from different tissues of coffee was optimized. Sufficiently high quality and quantity (225.6-454.8µg/g) of RNA was obtained by using the optimized protocol. The presence of two distinct bands of 28S rRNA and 18S rRNA in agarose gel proved the intactness of the RNA samples. The average spectrophotometric values of the isolated RNA ranged from 1.96 to 2.02 (A260/280) and 1.95 to 2.14 (A260/230), indicating the high quality of RNA devoid of polyphenols, polysaccharides and protein contamination. In the optimized protocol, addition of PVPP to the extraction buffer and a brief incubation of samples at 65°C and subsequent purification with potassium acetate resulted in good-quality RNA isolation. The suitability of RNA for downstream processing was confirmed by PCR amplification with cytochrome c oxidase gene-specific primers. The amplification of a single 392bp fragment using cDNA and 1.5kb fragment using genomic DNA samples confirmed the absence of DNA contamination. The present protocol is rapid and yielded good quality and quantity of RNA suitable for functional genomics studies.

English

Yam (Dioscorea species) and banana (Musa species) leaf sample contain high levels of polysaccharide and polyphenolic compounds. Extraction of good quality DNA from such leaves is usually problematic. Therefore, there is a need to extract good quality DNA in order to perform downstream DNA analysis, especially for the detection of viruses due to their low titration in some infected leaves. The cetyltrimethylammonium bromide (CTAB) DNA extraction method was selected in order to optimise DNA extraction from plant material with high level of polysaccharides. Four steps of the Lodhi et al. (1994) method were modified to be user friendly in less resource laboratories of developing countries. Yield and purity of the extracted DNAs were quantified using a NanoDrop 2000 spectrophotometer and by migration on an agarose gel after polymerase chain reaction (PCR) amplification. The results of the DNA yields and purity were in the range of 287.40±2.23 to 424.95±1.85 ng/ul and 2.10±0.05 to 2.19±0.04, respectively. Modification was found to yield DNA of reasonable quantity, quality and purity. Furthermore, the method can be used in a laboratory with less sophisticated waste disposal system. Key words: Deoxyribonucleic acid (DNA), cetyltrimethyl ammonium bromide (CTAB), yam, banana, tomato.

萌芽菊芋块茎对盐碱土壤胁迫的生理响应

PDF HTML阅读 XML下载 导出引用 引用提醒 萌芽菊芋块茎对盐碱土壤胁迫的生理响应 DOI: 10.5846/stxb201610262181 作者: 作者单位: 东北林业大学,东北林业大学盐碱地生物资源环境研究中心,东北林业大学,东北林业大学盐碱地生物资源环境研究中心 作者简介: 通讯作者: 中图分类号: 基金项目: 国家自然科学基金项目（31470467） Physiological response of sprouting Jerusalem artichoke tubers to saline-alkali stress Author: Affiliation: Northeast Forestry University,,Northeast Forestry University,Alkali Soil Natural Environmental Science Center,Northeast Forestry University Fund Project: 摘要 | 图/表 | 访问统计 | 参考文献 | 相似文献 | 引证文献 | 资源附件 | 文章评论 摘要:土壤盐碱化是影响全球农业生产和生态环境的重要问题。在农田、轻度盐碱草地和重度盐碱草地设置样地以块茎种植菊芋，次年5月块茎萌发阶段取块茎样品测定丙二醛、游离脯氨酸、可溶性糖含量以及抗氧化酶活性并进行蛋白质组学分析，分析了萌芽菊芋块茎对盐碱土壤胁迫的生理响应。0-20 cm土层的电导率（表征土壤可溶盐含量）表明从农田到轻度、重度盐碱草地土壤盐碱胁迫逐渐增强，丙二醛含量变化反映出菊芋块茎受害程度逐渐增加，并且基于游离脯氨酸的渗透调节能力也在逐渐增强。蛋白质组学分析结果显示与遗传信息加工相关的差异蛋白数量最多（占28.75%）且多为表达上调，意味着DNA复制和转录、蛋白质合成和折叠的相关蛋白在响应盐碱胁迫中发挥关键作用。碳水化合物及多糖代谢（占15%）、氨基酸代谢（占11.25%）以及能量代谢（占7.5%）相关的差异蛋白数量也较多，说明调节物质代谢平衡在萌芽菊芋块茎应对盐碱土壤胁迫过程中有重要作用。这些结果为揭示萌芽菊芋块茎适应盐胁迫的生理机制奠定了基础。 Abstract:Soil salinization is one of the most common abiotic stresses affecting plant growth and is becoming an important issue, owing to its impact on agricultural production and the environment. Saline-alkali soil is becoming particularly widespread and may cover more than 50% of all arable lands by the year 2050. The combination of soil salinization and high pH conditions represents a major impediment for plant growth and crop productivity. The study area was a typical alkalinized grassland in northeastern China. Jerusalem artichoke (Helianthus tuberosus) is an annual flowering plant that has been cultivated as a vegetable, fodder crop, and bioenergy material in many countries, owing to its high levels of polysaccharides, especially inulin. The ability to survive in the saline-alkali soils of semiarid areas is one of the most important characters of Jerusalem artichoke. Jerusalem artichoke tubers were sown in farmland, light saline-alkali, or severe saline-alkali soil and sprouting tubers were collected from the sample in May of the following year. The malondialdehyde, free proline, and soluble sugar contents, antioxidant enzyme activity, and protein profile were quantified, in order to assess the physiological response of Jerusalem artichoke to saline-alkali stress. The soil electrical conductivity (0-20 cm) indicated that the soluble salt contents of the three soil types were significantly different, with the lowest soluble salt content in the farmland soil and the greatest content in the severe saline-alkali soil. With increasing soluble salt content, the malondialdehyde content increased, obviously indicating higher levels of stress, and increased free proline content indicated that Jerusalem artichoke could enhance its resistance to salt stress via osmotic adjustment. To investigate the proteomic response of the sprouting tubers to saline-alkali stress, two-dimensional gel electrophoresis (2-DE) gels from three biological experiments were analyzed. Then, using ImageMaster 2D Platinum Software, we identified more than 1000 highly reproducible protein spots on the coomassie brilliant blue (CBB)-stained 2D gels. A total of 80 differentially expressed proteins were successfully identified using MALDI-TOF/TOF analysis, according to the peptide matching results provided by MASCOT. Among these proteins, 42 were detected in the sprouting H. tuberosus tubers from the light saline-alkali soil and 38 of the proteins accumulated differentially in the tubers grown in the severe saline-alkali soil. KEGG pathway analysis attributed these proteins to eleven different metabolic pathways, which included carbohydrate and polysaccharide metabolism (15%), energy metabolism (7.5%), genetic information processing (28.75%), amino acid metabolism (11.25%), nucleotide metabolism (2.5%), biosynthesis of secondary metabolites (3.75%), signal transduction (17.5%), transport and catabolism (2.5%), cell motility (2.5%), and unknown (8.75%). The differentially expressed proteins were mainly involved in genetic information processing, which might indicate that the regulation of proteins involved in DNA replication, transcription, protein synthesis, and protein folding are responsive to saline-alkali stress and play a pivotal role in salinity tolerance. Carbohydrate, energy, and amino acid metabolism-related proteins constituted one-third of the differentially expressed proteins, which suggests that metabolism homeostasis is important for the survival of seedlings exposed to saline-alkali stress. These findings provide new insight into the underlying molecular mechanisms of saline-alkali resistance in Jerusalem artichoke. 参考文献 相似文献 引证文献

Comparative Different DNA Isolation Protocols from Ziziphus spina-christi (L.) Leaves through RAPD and ISSR Markers

<p>Genomic analysis of plants relies on high quantity and quality of pure DNA. Extraction and purification of DNA from woody and medicinal plants, such as fruit trees present a great challenge due to accumulation of a large amount of co-purify with DNA, including polysaccharides, polyphenols and proteins. Therefore, it is necessary to optimize the extraction protocols to reduce these compounds to the lowest level. A study was conducted to compare six DNA extraction and precipitation methods for genomic analysis in<em> Ziziphus spina-christi</em> (L.) plant tissues. The results showed significant differences in DNA contents among the six methods. Quantity and quality of extracted genomic DNAs were compared by employing the spectrophotometer, Nano-Drop, agarose gel electrophoresis, digestion by restriction enzymes and polymerase chain reaction (PCR) methods and molecular marker such as RAPD and ISSR. The method of Vroh Bi et al., provided the best results (208.89 ng/μL) in terms of quantity and quality of DNA, and Doyle and Doyle method as second method for leaves sample were chosen. According to the results, the method of Bi et al. is recommended for DNA extraction from plant tissues having high level of polysaccharides and phenol compounds.</p>

Red bayberry extract prevents high-fat diet-induced metabolic disorders in C57BL/6 mice

Red bayberry, also called Chinese bayberry, is the fruit of Myrica rubra (Lour.) Sieb. et Zucc., which is used as both food and nutraceuticals in East Asian countries. The present study aimed to investigate whether the red bayberry extract (RBE) exhibits an anti-metabolic disorder effect on C57BL/6 mice fed with a high-fat diet (HFD). The extract contained high levels of polysaccharides and polyphenols. C57BL/6 mice were treated with a chow diet, HFD or HFD mixed with 1% (w/w) RBE for 8 weeks. Results showed that RBE blocked the body weight gain, improved glucose tolerance and insulin resistance, and lowered serum and liver triacylglycerol (TG) contents in HFD fed mice. Furthermore, RBE inhibited the mRNA expression levels of liver X receptor (LXR) α and its target genes, and increased peroxisome proliferator-activated receptor (PPAR) α and its target genes. The study suggested that RBE could prevent metabolic disorders in C57BL/6 mice induced by HFD, through the inhibition of the LXRα and the activation of PPARα signaling.

Effect of selected Saccharomyces cerevisiae yeast strains and different aging techniques on the polysaccharide and polyphenolic composition and sensorial characteristics of Cabernet Sauvignon red wines.

The objective of this work was to study the effect of two Saccharomyces cerevisiae yeast strains with different capabilities of polysaccharide liberation during alcoholic fermentation in addition to subsequent aging on lees with or without oak wood chips as well as aging with commercial inactive dry yeast on the physical, chemical and sensorial characteristics of Cabernet Sauvignon red wines. The HPS (high levels of polysaccharides) yeast strain released higher amounts of polysaccharides (429 g L(-1)) than EC1118 (390 g L(-1)) during alcoholic fermentation, but the concentration equalized during the aging period (424 and 417 g L(-1) respectively). All aging techniques increased the polysaccharide concentration, but the increase was dependent on the technique applied. A higher liberation of polysaccharides reduced the concentration of most of the phenolic families analyzed. Moreover, no clear effect of the different aging techniques used in this study on color stabilization was found. The HPS wines were better valued than the EC1118 wines by the panel of tasters after alcoholic fermentation. In general, the HPS wines showed better physicochemical and sensorial characteristics than the EC1118 wines. According to the results obtained during the aging period, all aging techniques contributed to improve wine quality, but it was difficult to establish the technique that allowed the best wine to be obtained, because it depended on the aging technique used and the period of aging.

English

Extraction of DNA in many plants is difficult because of the presence of metabolites that interfere with DNA isolation procedures and downstream applications such as DNA restriction, replications, amplification, as well as cloning. Modified procedure based on the hexadecyltrimethyl  ammoniumbromide (CTAB) method is used to isolate DNA from tissues containing high levels of polysaccharides. The procedure is applicable to both ripped and unripe fruits of Annona senegalensis . This modified CTAB (2%) protocol include the use of 1.4 M NaCl, 1% polyvinylpyrrolidine (PVP), 1% mercaptoethanol and 100% absolute ethanol in the extraction as well as reducing the centrifugation times during the separation and precipitation of the DNA. This method solved the problems of DNA contamination, degradation and low yield due to binding or co-precipitation with starches. The isolated DNA proved amenable to polymerase chain reaction (PCR) amplification and restriction digestion. This technique is fast, reproducible, and can be applied for simple sequence repeats (SSR)-PCR markers identification. Key words : Annona senegalensis , genomic DNA, fruits, modified, markers.

Chapter Five - Extracellular Polysaccharides Produced by Marine Bacteria

Extracellular polysaccharides (EPSs) produced by microorganisms are a complex mixture of biopolymers primarily consisting of polysaccharides, as well as proteins, nucleic acids, lipids, and humic substances. Microbial polysaccharides are multifunctional and can be divided into intracellular polysaccharides, structural polysaccharides, and extracellular polysaccharides or exopolysaccharides. Recent advances in biological techniques allow high levels of polysaccharides of interest to be produced in vitro. Biotechnology is a powerful tool to obtain polysaccharides from a variety of marine microorganisms, by controlling the growth conditions in a bioreactor while tailoring the production of biologically active compounds. The aim of this chapter is to give an overview of current knowledge on extracellular polysaccharides producing marine bacteria isolated from marine environment.

A QUICK, EFFICIENT, AND COST-EFFECTIVE METHOD FOR ISOLATING HIGH-QUALITY TOTAL RNA FROM TOMATO FRUITS, SUITABLE FOR MOLECULAR BIOLOGY STUDIES

Tomato (Solanum lycopersicum L.) is the primary model for the study of fleshy fruits, and research on this species has elucidated many aspects of fruit physiology, development, and metabolism. However, for advancing such studies at molecular biology levels, the RNA isolation from fruit tissues is often essential. The RNA isolation from tomato fruits is complicated because of the presence of high levels of polysaccharides, polyphenolics, pigments, and secondary metabolites and also the varying water content during development. Here, we present an optimized protocol for the isolation of total RNA from the fruit tissues at different developmental stages. In comparison to the previous methods described for the RNA isolation from tomato fruit, this method has the advantages that it does not involve the use of guanidine salts, lyophilizers, and commercial reagents, reduces the time and cost of extraction, overcomes the high water content problem, and promotes RNA quality by inhibiting RNA degradation and minimizing the gDNA, polyphenolic and polysaccharide contaminations. Using this method, high yields of high-purity and intact RNA samples were obtained as confirmed by the spectrophotometric readings and the electrophoresis on denaturing agarose gels. The isolated RNA was employed as a robust template for cDNA synthesis, reverse transcriptase-polymerase chain reaction (RT-PCR), and temporal gene expression analysis. The functionality of the isolated RNA was further demonstrated through cloning full-length cDNAs encoding β-galactosidase proteins by RT-PCR and sequencing.

Modified method for combined DNA and RNA isolation from peanut and other oil seeds

Isolation of good quality RNA and DNA from seeds is difficult due to high levels of polysaccharides, polyphenols, and lipids that can degrade or co-precipitate with nucleic acids. Standard RNA extraction methods utilizing guanidinium-phenol-chloroform extraction has not shown to be successful. RNA isolation from plant seeds is a prerequisite for many seed specific gene expression studies and DNA is necessary in marker-assisted selection and other genetic studies. We describe a modified method to isolate both RNA and DNA from the same seed tissue and have been successful with several oil seeds including peanut, soybean, sunflower, canola, and oil radish. An additional LiCl precipitation step was added to isolate both RNA and DNA from the same seed tissues. High quality nucleic acids were observed based on A(260)/A(280) and A(260)/A(230) ratios above 2.0 and distinct bands on gel-electrophoresis. RNA was shown to be suitable for reverse transcriptase polymerase chain reaction based on actin or 60S ribosomal primer amplification and DNA was shown to have a single band on gel-electrophoresis analysis. This result shows that RNA and DNA isolated using this method can be appropriate for molecular studies in peanut and other oil containing seeds.