A rapid and efficient method for isolation of total RNA from Euglena gracilis (Euglenoidea)

González-Mendoza, Daniel; Morales, Adriana; Brito-Vera, H

doi:10.4238/vol8-2gmr590

Cited by 5 publications

(5 citation statements)

References 5 publications

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“…Total RNA was extracted as described (González-Mendoza et al, 2009) after Nycodenz gradient density separation of the bacterial community for 15 min at 4 1C from sediments stored at À80 1C. Total RNA was purified using the RNEasy Mini kit (Qiagen, Valencia, CA, USA) and digested with DNAse (Fermentas, Glen Burnie, MD, USA).…”

Section: Pcr and Reverse Transcriptionmentioning

confidence: 99%

Metabolic diversity among main microorganisms inside an arsenic-rich ecosystem revealed by meta- and proteo-genomics

et al. 2011

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By their metabolic activities, microorganisms have a crucial role in the biogeochemical cycles of elements. The complete understanding of these processes requires, however, the deciphering of both the structure and the function, including synecologic interactions, of microbial communities. Using a metagenomic approach, we demonstrated here that an acid mine drainage highly contaminated with arsenic is dominated by seven bacterial strains whose genomes were reconstructed. Five of them represent yet uncultivated bacteria and include two strains belonging to a novel bacterial phylum present in some similar ecosystems, and which was named 'Candidatus Fodinabacter communificans.' Metaproteomic data unravelled several microbial capabilities expressed in situ, such as iron, sulfur and arsenic oxidation that are key mechanisms in biomineralization, or organic nutrient, amino acid and vitamin metabolism involved in synthrophic associations. A statistical analysis of genomic and proteomic data and reverse transcriptase-PCR experiments allowed us to build an integrated model of the metabolic interactions that may be of prime importance in the natural attenuation of such anthropized ecosystems.

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Section: Pcr and Reverse Transcriptionmentioning

confidence: 99%

Metabolic diversity among main microorganisms inside an arsenic-rich ecosystem revealed by meta- and proteo-genomics

et al. 2011

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“…High levels of oxidised phenolics, similar to those found in avocado, are well known to hinder DNA and RNA extraction [ 16 , 17 , 18 , 19 , 20 , 21 , 22 ]. While phenol:C:IAA (phenol:chloroform:isoamyl alcohol) is commonly used during CTAB extraction, the DNA degrades in the case of avocado.…”

Section: Resultsmentioning

confidence: 99%

“…Many have ionic similarity or compatibility to nucleic acids; thus, they co-precipitate [ 14 , 15 ], whereas others react with various compounds during the process. Phenolic compounds, which are oxidised after cell lysis, allowing them to bind to nucleic acids irreversibly, co-precipitate and make nucleic acids challenging to dissolve [ 16 , 17 , 18 , 19 , 20 , 21 , 22 ]. Contaminants can also impact yield and the accurate quantification of nucleic acids and can inhibit enzymes in downstream applications, including PCR and library preparation processes for nucleic acid sequencing.…”

Section: Introductionmentioning

confidence: 99%

A Comprehensive High-Quality DNA and RNA Extraction Protocol for a Range of Cultivars and Tissue Types of the Woody Crop Avocado

Nath

Fletcher

Hayward

et al. 2022

Plants

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High-quality DNA and RNA forms the basis of genomic and genetic investigations. The extraction of DNA and RNA from woody trees, like avocado (Persea americana Mill.), is challenging due to compounds which interact with nucleic acids and influence separation. Previously reported methods of DNA and RNA extraction from avocado have issues of low yield, quality and applicability across different cultivars and tissue types. In the current study, methods have been optimised for high-quality DNA extraction from 40 avocado cultivars and RNA extraction from multiple tissue types, including roots, stem, leaves, flowers and fruits. The method is based on the modification of the cetyltrimethylammonium bromide buffer, centred around the specific optimisation of chemicals, such as sodium dodecyl sulphate, polyvinylpyrrolidone, sodium sulphite, polyethylene glycol and β-mercaptoethanol. The DNA extraction method yielded high-molecular weight DNA from the leaf tissue of 40 avocado cultivars belonging to Mexican, Guatemalan and West Indian avocado horticultural groups. The method was further optimised for RNA extraction from different avocado plant parts, enabling extraction using amounts as low as ~10 mg of starting material. The DNA and RNA extracted was successfully used for long- and short-read sequencing and gene expression analysis. The methods developed may also be applicable to other recalcitrant plant species.

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“…Total RNA were extracted according to González‐Mendoza and colleagues () from 10 independent cultures of E. mutabilis and E. gracilis cultivated either in the absence or in the presence of arsenite (AsIII, 300 ppm). The 10 total RNA extracts of each of the four conditions ( E. gracilis with AsIII, E. gracilis without AsIII, E. mutabilis with AsIII, E. mutabilis without AsIII) were pooled in order to obtain four different total RNA extracts.…”

Section: Methodsmentioning

confidence: 99%

Arsenic hypertolerance in the protist Euglena mutabilis is mediated by specific transporters and functional integrity maintenance mechanisms

Halter

Andrés

Plewniak

et al. 2014

Environmental Microbiology

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Arsenic is a toxic metalloid known to cause multiple and severe cellular damages, including lipid peroxidation, protein misfolding, mutagenesis and double and single-stranded DNA breaks. Thus, exposure to this compound is lethal for most organisms but some species such as the photosynthetic protist Euglena mutabilis are able to cope with very high concentrations of this metalloid. Our comparative transcriptomic approaches performed on both an arsenic hypertolerant protist, i.e. E. mutabilis, and a more sensitive one, i.e. E. gracilis, revealed multiple mechanisms involved in arsenic tolerance. Indeed, E. mutabilis prevents efficiently the accumulation of arsenic in the cell through the expression of several transporters. More surprisingly, this protist induced the expression of active DNA reparation and protein turnover mechanisms, which allow E. mutabilis to maintain functional integrity of the cell under challenging conditions. Our observations suggest that this protist has acquired specific functions regarding arsenic and has developed an original metabolism to cope with acid mine drainages-related stresses.

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A rapid and efficient method for isolation of total RNA from Euglena gracilis (Euglenoidea)

Cited by 5 publications

References 5 publications

Metabolic diversity among main microorganisms inside an arsenic-rich ecosystem revealed by meta- and proteo-genomics

Metabolic diversity among main microorganisms inside an arsenic-rich ecosystem revealed by meta- and proteo-genomics

A Comprehensive High-Quality DNA and RNA Extraction Protocol for a Range of Cultivars and Tissue Types of the Woody Crop Avocado

Arsenic hypertolerance in the protist Euglena mutabilis is mediated by specific transporters and functional integrity maintenance mechanisms

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