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Chemical lysis of cyanobacteria.

Mehta KK, Evitt NH, Swartz JR - J Biol Eng (2015)

Bottom Line: We have developed a mixture of enzymes and chemicals that completely lyse cyanobacteria.Since the treatment involves only readily-available chemicals and simple proteins that degrade the components of the cyanobacterial cell wall, it can easily be used in high-throughput applications requiring lysis for subsequent intracellular measurements.Chemical lysis can be superior to existing techniques because of its convenience, reliability, and amenability to a variety of downstream applications.

View Article: PubMed Central - PubMed

Affiliation: Department of Bioengineering, Stanford University, Stanford, CA 94305 USA.

ABSTRACT
We have developed a mixture of enzymes and chemicals that completely lyse cyanobacteria. Since the treatment involves only readily-available chemicals and simple proteins that degrade the components of the cyanobacterial cell wall, it can easily be used in high-throughput applications requiring lysis for subsequent intracellular measurements. Our lysis technique consistently enables complete lysis of several different cyanobacterial strains, and we demonstrated that DNA, mRNA, and proteins are preserved in the lysates. Chemical lysis can be superior to existing techniques because of its convenience, reliability, and amenability to a variety of downstream applications.

No MeSH data available.


Related in: MedlinePlus

Comparison of cyanophage lysozyme with T4 lysozyme for cyanobacterial lysis. The concentration of cyanophage lysozyme (purified in-house) and T4 lysozyme (MCLab) was titrated to compare their effectiveness in lysing Synechocystis sp. PCC 6803. Each reaction contained lysozyme at the indicated concentration, 25 mM MES buffer (pH 6.2), and 1 X BugBuster reagent. Reactions were incubated for 90 min at 37 °C. We estimated that the maximum lysis achieved here corresponded to 50 % of total lysis; approx. 25 % of the cells were lysed with BugBuster alone. Error bars are standard deviations of triplicate reactions
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Fig2: Comparison of cyanophage lysozyme with T4 lysozyme for cyanobacterial lysis. The concentration of cyanophage lysozyme (purified in-house) and T4 lysozyme (MCLab) was titrated to compare their effectiveness in lysing Synechocystis sp. PCC 6803. Each reaction contained lysozyme at the indicated concentration, 25 mM MES buffer (pH 6.2), and 1 X BugBuster reagent. Reactions were incubated for 90 min at 37 °C. We estimated that the maximum lysis achieved here corresponded to 50 % of total lysis; approx. 25 % of the cells were lysed with BugBuster alone. Error bars are standard deviations of triplicate reactions

Mentions: A primary question was whether the new cyanophage lysozyme would perform better than other lysozymes against cyanobacteria. We tested this by comparing cyanophage lysozyme activity on Synechocystis sp. PCC 6803 to that of T4 lysozyme, a commercially available enzyme commonly used for E. coli lysis (Fig. 2). Overall, we found that cyanophage lysozyme was equivalent to T4 lysozyme in promoting cyanobacterial lysis (although at low lysozyme concentrations there was a slight advantage to using T4 lysozyme). Based on this, most subsequent experiments were done with T4 lysozyme.Fig. 2


Chemical lysis of cyanobacteria.

Mehta KK, Evitt NH, Swartz JR - J Biol Eng (2015)

Comparison of cyanophage lysozyme with T4 lysozyme for cyanobacterial lysis. The concentration of cyanophage lysozyme (purified in-house) and T4 lysozyme (MCLab) was titrated to compare their effectiveness in lysing Synechocystis sp. PCC 6803. Each reaction contained lysozyme at the indicated concentration, 25 mM MES buffer (pH 6.2), and 1 X BugBuster reagent. Reactions were incubated for 90 min at 37 °C. We estimated that the maximum lysis achieved here corresponded to 50 % of total lysis; approx. 25 % of the cells were lysed with BugBuster alone. Error bars are standard deviations of triplicate reactions
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4478636&req=5

Fig2: Comparison of cyanophage lysozyme with T4 lysozyme for cyanobacterial lysis. The concentration of cyanophage lysozyme (purified in-house) and T4 lysozyme (MCLab) was titrated to compare their effectiveness in lysing Synechocystis sp. PCC 6803. Each reaction contained lysozyme at the indicated concentration, 25 mM MES buffer (pH 6.2), and 1 X BugBuster reagent. Reactions were incubated for 90 min at 37 °C. We estimated that the maximum lysis achieved here corresponded to 50 % of total lysis; approx. 25 % of the cells were lysed with BugBuster alone. Error bars are standard deviations of triplicate reactions
Mentions: A primary question was whether the new cyanophage lysozyme would perform better than other lysozymes against cyanobacteria. We tested this by comparing cyanophage lysozyme activity on Synechocystis sp. PCC 6803 to that of T4 lysozyme, a commercially available enzyme commonly used for E. coli lysis (Fig. 2). Overall, we found that cyanophage lysozyme was equivalent to T4 lysozyme in promoting cyanobacterial lysis (although at low lysozyme concentrations there was a slight advantage to using T4 lysozyme). Based on this, most subsequent experiments were done with T4 lysozyme.Fig. 2

Bottom Line: We have developed a mixture of enzymes and chemicals that completely lyse cyanobacteria.Since the treatment involves only readily-available chemicals and simple proteins that degrade the components of the cyanobacterial cell wall, it can easily be used in high-throughput applications requiring lysis for subsequent intracellular measurements.Chemical lysis can be superior to existing techniques because of its convenience, reliability, and amenability to a variety of downstream applications.

View Article: PubMed Central - PubMed

Affiliation: Department of Bioengineering, Stanford University, Stanford, CA 94305 USA.

ABSTRACT
We have developed a mixture of enzymes and chemicals that completely lyse cyanobacteria. Since the treatment involves only readily-available chemicals and simple proteins that degrade the components of the cyanobacterial cell wall, it can easily be used in high-throughput applications requiring lysis for subsequent intracellular measurements. Our lysis technique consistently enables complete lysis of several different cyanobacterial strains, and we demonstrated that DNA, mRNA, and proteins are preserved in the lysates. Chemical lysis can be superior to existing techniques because of its convenience, reliability, and amenability to a variety of downstream applications.

No MeSH data available.


Related in: MedlinePlus