A rapid and cost-effective method of genomic DNA isolation from cyanobacterial culture, mat and soil suitable for genomic fingerprinting and community analysis

Srivastava, Amrita; Ara, Anjum; Bhargava, Poonam; Mishra, Yogesh; P., S.; C., L.

doi:10.1007/s10811-006-9144-5

Cited by 42 publications

(25 citation statements)

References 24 publications

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“…The sonic shock treatment in an ultrasonic cleaner bath and homogenization using sterile glass beads on a vortex mixer was used for clump forming cyanobacteria to separate and break the filaments (Fiorea et al 2000;Srivastava et al 2007). However, our method does not require such treatments as we were able to extract genomic DNA efficiently from the Rivularia sp.…”

Section: Resultsmentioning

confidence: 99%

“…This method also skips the initial washing of the cells with washing buffer that has been used by several workers to remove the extracellular polysaccharides (Margheri et al 1999;Fiorea et al 2000;Srivastava et al 2007;Morin et al 2010). Sarkosyl (0.1%) has also been used to remove the lipid polysaccharide which was essential for cell lysis by lysozyme (Wu et al 2000), however, all these additional steps will render DNA fragmentation by longer exposure to deoxyribonucleases and integrity of the genomic DNA will be compromised.…”

Section: Resultsmentioning

confidence: 99%

“…However, most of the filamentous cyanobacteria are often surrounded by a sheath which acts as a barrier for the enzymatic action of lysozyme on the cell wall (Philippis and Vincenzini 1998). Alternatively, sonication of samples was used to disrupt the cells (Sinha et al 2001;Srivastava et al 2007), however, sonicators generate sound waves that are outside the normal range of hearing and can cause hearing damage. It is also time taking and there is also chance of contamination of samples with sonication when working with several samples, additionally, DNA molecules are always susceptible to fragmentation with sonication.…”

Section: Introductionmentioning

confidence: 99%

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An improved method for genomic DNA extraction from cyanobacteria

Singh

Rastogi

Häder

et al. 2010

World J Microbiol Biotechnol

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We present an improved method for genomic DNA extraction from cyanobacteria by updating the earlier method from our group (Sinha et al. 2001) that does not require lysozyme treatment or sonication to lyse the cells. This method use lysis buffer to lyse the cells and also skips the initial treatments to remove the exopolysaccharides or to break the clumps. To test the efficacy of the method DNA was extracted from the freshwater cyanobacteria Anabaena variabilis PCC 7937, Anabaena sp. PCC 7120, Synechocystis sp. PCC 6803, Synechococcus sp. PCC 6301 and Rivularia sp. HKAR-4 (Accession number: FJ939128). The spectrophotometric and gel electrophoresis analysis revealed high yield and high quality of genomic DNA extracted by this method. Furthermore, the RAPD resulted in the amplification of unidentified genomic regions of various lengths; however, rDNA amplification gave only one band of 1.5 kb in all studied cyanobacteria. Thymine dimer detection study revealed that thymine dimers are induced only by UV-B radiation in A. variabilis PCC 7937 and there is no effect of PAR and UV-A on its genome.Collectively, all these findings put forward the applicability of this method in different studies and purposes.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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An improved method for genomic DNA extraction from cyanobacteria

Singh

Rastogi

Häder

et al. 2010

World J Microbiol Biotechnol

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“…Genomic DNA was isolated using the method described by Srivastava et al (2007). PCR was performed in a 20 ml reaction mixture containing 100 ng DNA, PCR buffer, 4.5 mM MgCl 2 , 200 mM dNTPs, 10 pmol of each primer and 0.2 U Taq DNA polymerase.…”

Section: Methodsmentioning

confidence: 99%

Assessment of salinity-induced photorespiratory glycolate metabolism in Anabaena sp. PCC 7120

et al. 2011

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This paper reports an investigation of salinity-induced glycolate metabolism in the cyanobacterium Anabaena sp. PCC 7120 (hereafter Anabaena PCC 7120). Quantitative analysis of transcripts for the photosynthesis-associated genes encoding ribulose-1,5-bisphosphate carboxylase oxygenase (Rubisco), phosphoribulokinase and transketolase, as well as those involved in glycolate metabolism (phosphoglycolate phosphatase, glycolate oxidase, alanine-glyoxylate aminotransferase and serine hydroxymethyltransferase) was performed. The expression of all investigated photosynthesis-associated genes except Rubisco was downregulated after 24 h NaCl treatment. However, under the same conditions, the transcripts encoding enzymes involved in glycolate metabolism were overexpressed. This was further confirmed by the quantitative analysis of the intermediates involved in glycolate metabolism. The intracellular levels of organic acids (glyceric, glycolic and glyoxylic acids) and amino acids (glycine and serine) were elevated in salt-treated cells as compared to those in the control cells. Transcriptional inhibition of photosynthesis-associated genes, and upregulation of genes and enhanced synthesis of intermediates associated with glycolate metabolism, indicate the occurrence of this photorespiratory metabolic pathway metabolism in Anabaena PCC 7120 under salt stress. INTRODUCTIONThe evolution of cyanobacteria over 2.7 billion years significantly contributed to a major transition in the history of life on Earth (Buick, 1992). Cyanobacterial photosynthesis resulted in a tremendous increase in the oxygen concentration of the atmosphere and the emergence of oxygen-dependent life. Due to their ability to adapt frequently to changing environments, cyanobacteria currently occupy most ecological niches on our planet.An important habitat for cyanobacteria is rice fields, where they significantly contribute to the availability of nitrogen for the crop (Singh, 1961). The rice agro-ecosystem, including cyanobacteria, is subjected to several abiotic stresses, including salt stress caused by anthropogenic activities (Srivastava et al., 2009). It has been reported that high salinity not only inhibits photosynthesis and hence the carbon pool of freshwater cyanobacteria but also decreases the amount of fixed carbon available for the synthesis of compatible solutes (Ferjani et al., 2003). Responses of freshwater cyanobacteria to salt stress have been studied by several research groups at the genomic, transcriptomic and proteomic levels Huang et al., 2006;Kanesaki et al., 2002; Marin et al., 2004;Srivastava et al., 2008). These studies reported salt stress responses of several cyanobacterial species in relation to specific genes encoding stress proteins, photosynthetic proteins and proteins involved in the synthesis of compatible solutes. However, salinity-induced changes in metabolism related to glycolate metabolism (photorespiration) have been little studied in cyanobacteria.Photorespiration evolved as an essential disadvantage of oxygenic life, as...

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“…The complicated components of soils, especially humic acid, phenolic compound, heavy metal ion lead to difficulties in the process of DNA extraction. The humic acid, as a three-dimensional structure, can bind other compounds and absorb water, ions and organic molecules [16,20]. Because its physicochemical property is similar to that of nucleic acids, it can be co-extracted with genomic DNA from soil samples and it is hard to seperate from each other.…”

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confidence: 99%

A developed DNA extraction method for different soil samples

Liu

Yan

et al. 2010

J. Basic Microbiol.

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Four DNA extraction methods namely SDS-hyperhaline method (I), modified SDS-hyperhaline method (II), indirect method (III), alkaline lysis method (IV) were evaluated by comparing DNA yield, spectrophotometric quality, genomic integrity and PCR suitability in this paper. The results showed that high DNA yields were obtained by method I, II and IV. However, higher quality of DNA was gained by method III and IV. Based on the results of the Pulsed-Field Gel Electrophoresis (PFGE), the completeness of DNA extracted by method IV was the best. About 6.0 microg DNA can be recovered from 1.0 g soil by method IV which involved to lysis cell by SDS and to precipitate impurities by adding potassium acetate and magnesium chloride Therefore, it is confirmed that method IV is a novel, reliable and versatile method for large-scale DNA extraction involving less purification steps for various soil samples.

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A rapid and cost-effective method of genomic DNA isolation from cyanobacterial culture, mat and soil suitable for genomic fingerprinting and community analysis

Cited by 42 publications

References 24 publications

An improved method for genomic DNA extraction from cyanobacteria

An improved method for genomic DNA extraction from cyanobacteria

Assessment of salinity-induced photorespiratory glycolate metabolism in Anabaena sp. PCC 7120

A developed DNA extraction method for different soil samples

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