Single-cell total RNA extraction from marine protists (e.g. Acantharia, Strombidium cf basimorphum, and Prymnesium parvum) v1 (protocols.io.bp6xmrfn)

protocols.io ◽  
2020 ◽  
Author(s):  
Joost Mansour ◽  
Konstantinos Anestis ◽  
Fabrice Not ◽  
Uwe John
Keyword(s):  
Single Cell ◽  
Rna Extraction ◽  
Prymnesium Parvum ◽  
Total Rna ◽  
Marine Protists ◽  
Total Rna Extraction

cDNA library preparation from total RNA extracts of Single-cell marine protists (e.g. Acantharia, Strombidium basimorphum, and Prymnesium parvum) for transcriptome sequencing v2

2022 ◽  
Author(s):  
Joost S S. Mansour ◽  
Konstantinos Anestis ◽  
Fabrice Not ◽  
Uwe John
Keyword(s):  
Single Cell ◽  
Cdna Library ◽  
Mammalian Cells ◽  
Single Cells ◽  
Rna Extraction ◽  
Cdna Libraries ◽  
Prymnesium Parvum ◽  
Large Variability ◽  
Total Rna ◽  
Marine Protists

Many marine protists are not culturable and therefore challenging to study, nonetheless, they are essential in all marine ecosystems. The development of single-cell techniques is allowing for more marine protists to be studied. Such genomic approaches aim to help to disentangle heterotrophic processes such as phagotrophy from osmotrophy and phototrophic-induced anabolic activities. This information will then support cellular and metabolic modeling by better elucidating the physiological mechanisms and quantifying their importance in different scenarios. However, single-cell protocols and low input RNA kits for transcriptomics are usually made for and tested with mammalian cells, as such the feasibility and efficiency of single-cell transcriptomics on highly diverse mixotrophic protists is not always known. Often single-cell transcriptomics of microbial eukaryotes shows low transcript recovery rates and large variability. We report on transcriptomic methods that we have successfully performed on single cells of Acantharia, Strombidium basimorphum, and Prymnesium parvum. This protocol follows up after total RNA extraction (from the protocol at dx.doi.org/10.17504/protocols.io.bp6xmrfn) to prepare cDNA libraries for Illumina sequencing. The described protocol uses the SMART-Seq4 kit (Takara #634891) for cDNA synthesis and amplification, but this can also be successfully performed with the NEBNext kit (NEB #E6421). The NEBNext kit protocol is very similar to the protocol described here and generally the manufacture's protocol can be followed but see the notes at step 4 and step 18 of this protocol, and do the final elution after cDNA purification in 10 mM Tris (pH 8.0). The subsequent cDNA library is prepared following the .

Total RNA Extraction from Caenorhabditis elegans

2020 ◽  
Vol 2020 (9) ◽  
pp. pdb.prot101683
Author(s):  
Michael R. Green ◽  
Joseph Sambrook
Keyword(s):  
Caenorhabditis Elegans ◽  
Rna Extraction ◽  
Total Rna ◽  
Total Rna Extraction

A RAPID AND EFFECTIVE METHOD FOR TOTAL RNA EXTRACTION FROM CONIDIA AND MYCELIUM OF SEPTORIA TRITICI

2003 ◽  
Vol 11 (1) ◽  
pp. 53-59 ◽  
Author(s):  
JIAN-RONG GUO ◽  
KAMEL A. ABD-ELSALAM ◽  
FRANK SCHNIEDER ◽  
JOSEPH-ALEXANDER VERREET
Keyword(s):  
Rna Extraction ◽  
Septoria Tritici ◽  
Total Rna ◽  
Total Rna Extraction

scholarly journals Comparison of different methods for total RNA extraction from sclerotia of Rhizoctonia solani

2014 ◽  
Vol 17 (1) ◽  
pp. 50-54 ◽  
Author(s):  
Canwei Shu ◽  
Si Sun ◽  
Jieling Chen ◽  
Jianyi Chen ◽  
Erxun Zhou
Keyword(s):  
Rhizoctonia Solani ◽  
Rna Extraction ◽  
Total Rna ◽  
Total Rna Extraction

scholarly journals Candida albicans Culture, Cell Harvesting, and Total RNA Extraction

BIO-PROTOCOL ◽  
2020 ◽  
Vol 10 (21) ◽  
Author(s):  
Max Cravener ◽  
Aaron Mitchell
Keyword(s):  
Candida Albicans ◽  
Rna Extraction ◽  
Culture Cell ◽  
Total Rna ◽  
Cell Harvesting ◽  
Total Rna Extraction

scholarly journals An efficient method for total RNA extraction from leaves of arboreal species from the Brazilian Cerrado

Rodriguésia ◽  
2020 ◽  
Vol 71 ◽  
Author(s):  
Geisiane Alves Rocha ◽  
Vanessa Duarte Dias ◽  
Renato Carrer-Filho ◽  
Marcos Gomes da Cunha ◽  
Érico de Campos Dianese
Keyword(s):  
Native Species ◽  
Rna Extraction ◽  
Woody Species ◽  
Extraction Methods ◽  
Native Plant ◽  
Brazilian Cerrado ◽  
Native Plant Species ◽  
Total Rna ◽  
Ctab Method ◽  
Total Rna Extraction

Abstract Considering the lack of information on RNA extraction from arboreal species, specially from the Brazilian Cerrado, the aim of this study was to test RNA extraction methods for a wide variety of native plant species from this biome. The methods tested consisted of: (i) TRIzol® reagent, (ii) TRIzol® reagent with modifications, (iii) CTAB buffer, and (iv) Modified CTAB buffer, initially for leaf samples of Xylopia aromatica and Piper arboreum. Later the procedure with the best results was used to obtain purified RNA from 17 other native species. Based on A260/A280 absorbance ratio the Modified CTAB method was the best for total RNA extraction for those woody species. Ten out of eleven species tested through RT-PCR generated fragments of the expected size from the total RNA extracted by the selected method, confirming it as the best option to obtain high-quality RNA for molecular analyses and for use in the detection of viruses infecting these tree species.

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