A high throughput assay of lichenase activity with Congo red dye in plants
| Autor: | Viktoriia A. Fridman, Aleksandra V. Suhorukova, O. S. Pavlenko, Igor V. Deineko, I. V. Goldenkova-Pavlova, A. A. Tyurin |
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| Jazyk: | angličtina |
| Rok vydání: | 2021 |
| Předmět: |
Plant reporter
QH301-705.5 Expression Plant Science Congo red Reporter proteins SB1-1110 chemistry.chemical_compound Quantitative expression Gene expression Expression analysis Genetics Biology (General) Lichenase biology Chemistry Methodology Direct observation Plant culture biology.organism_classification Biochemistry Lichenase activity Regulatory sequence Clostridium thermocellum Quantitative analysis (chemistry) Biotechnology |
| Zdroj: | Plant Methods, Vol 17, Iss 1, Pp 1-11 (2021) Plant Methods |
| ISSN: | 1746-4811 |
| Popis: | Background Since the beginning of the use of reporter proteins for expression analysis, a variety of approaches have been developed and proposed; both qualitative and quantitative. The lack of simple methods for direct observation of gene expression in living organisms makes it necessary to continue to propose new methods. In this work, we consider a method for the quantitative analysis of the expression of thermostable lichenase from Clostridium thermocellum used as a sensitive reporter protein. Results In this study, we report the design a high throughput fluorometric method for quantification of thermostable lichenase C. thermocellum using Congo red and further experimental verification of its relevance and efficiency in assessment of the functional role of regulatory sequences in the plant cell. Conclusions The specific interaction between the dye Congo red and $$\beta$$ β -d-glucans formed the background for designing a high-throughput fluorometric assay for quantification of C. thermocellum thermostable lichenase as a reporter protein for plants. This assay (i) makes it possible to precisely measure the amount of reporter protein in a plant sample; (ii) has shown a high sensitivity for quantification of thermostable lichenase; (iii) is more time- and cost-efficient as compared with the Somogyi–Nelson assay; and (iv) is to the least degree dependent on the presence of the tested buffer components as compared with the Somogyi–Nelson assay. |
| Databáze: | OpenAIRE |
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