Optimization of transformation conditions of the yeast Saccharomyces cerevisiae to determine coregulators of the transcription factor NIN in a yeast two-hybrid system
- Authors: Dymo A.1, Kantsurova E.S.1, Dolgikh A.V.1,2, Dolgikh E.A.1
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Affiliations:
- All-Russia Research Institute for Agricultural Microbiology
- Saint Petersburg State University
- Section: Methodology in ecological genetics
- Submitted: 15.01.2024
- Accepted: 01.05.2024
- Published: 29.05.2024
- URL: https://journals.eco-vector.com/ecolgenet/article/view/625670
- DOI: https://doi.org/10.17816/ecogen625670
- ID: 625670
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Abstract
The method of analyzing protein-protein interactions using the yeast two-hybrid system in Saccharomyces cerevisiae cells is actively used in the search for protein co-regulators. Advantages of this approach include the synthesis of proteins within cells in vivo, the relative speed of this analysis, and high specificity. This method also allows for large-scale screening of libraries of cloned fragments of complementary DNA (cDNA) that are translated within cells. A key factor in the success of such screens is the efficiency of yeast cell transformation, as the resolution of the analysis is based on this. As part of our research, we identified factors that influence the effectiveness of transformation. In particular, we showed that one of the key factors influencing transformation efficiency is the molecular weight of the polyethylene glycol used for chemical transformation of yeast cells and the number of cell division cycles that the cell culture undergoes. We also investigated the effect of the amount and size of plasmids used during transformation. Based on the obtained data, the optimal parameters to achieve a high level of competency in yeast cells were determined. Using an optimized transformation protocol, we screened a cDNA library to explore possible co-regulators of the NIN transcription factor which is a key regulator of the development of the legume-rhizobia symbiosis.
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About the authors
Alina Dymo
All-Russia Research Institute for Agricultural Microbiology
Email: dymoalina@yandex.ru
ORCID iD: 0000-0002-5919-2487
ResearcherId: AAF-3244-2021
младший научный сотрудник
Russian FederationElizaveta S. Kantsurova
All-Russia Research Institute for Agricultural Microbiology
Email: rudaya.s.e@gmail.com
ORCID iD: 0000-0002-3081-9880
SPIN-code: 4752-1910
Junior Researcher, Signal Regulation Laboratory
Russian Federation, Saint PetersburgAlexandra Vyacheslavovna Dolgikh
All-Russia Research Institute for Agricultural Microbiology; Saint Petersburg State University
Email: sqshadol@gmail.com
ORCID iD: 0000-0003-1845-9701
Scopus Author ID: 5719038282
ResearcherId: ABC-2930-2020
engineer
Russian Federation, 3 Podbelsky chausse, Pushkin, Saint Petersburg, 196608; Saint PetersburgElena A. Dolgikh
All-Russia Research Institute for Agricultural Microbiology
Author for correspondence.
Email: dol2helen@yahoo.com
ORCID iD: 0000-0002-5375-0943
SPIN-code: 4453-2060
Scopus Author ID: 6603496335
ResearcherId: G-6363-2017
Cand. Sci. (Biol.), Leading Researcher, Laboratory of Molecular and Cellular Biology
Russian Federation, Podbelsky chausse 3, 196608, St.-Petersburg, RussiaReferences
- Berggård T., Linse S., James P. Methods for the detection and analysis of protein–protein interactions. PROTEOMICS. 2007;7(16):2833–2842. doi: 10.1002/pmic.200700131
- Zhou M., Li Q., Wang R. Current Experimental Methods for Characterizing Protein–Protein Interactions. ChemMedChem. 2016;11(8):738–756. doi: 10.1002/cmdc.201500495
- Cuéllar A.P. et al. Yeast Two-Hybrid Analysis of Jasmonate Signaling Proteins. Jasmonate Signaling / ed. Goossens A., Pauwels L. Totowa, NJ: Humana Press. 2013;1011:173–185. doi: 10.1007/978-1-62703-414-2_14
- Caufield J.H., Sakhawalkar N., Uetz P. A comparison and optimization of yeast two-hybrid systems. Methods. 2012;58(4):317–324. doi: 10.1016/j.ymeth.2012.12.001
- Yang F. et al. Development and application of a recombination-based library versus library high- throughput yeast two-hybrid (RLL-Y2H) screening system. Nucleic Acids Res. 2018;46(3):e17–e17. doi: 10.1093/nar/gkx1173
- Erffelinck M.-L. et al. A user-friendly platform for yeast two-hybrid library screening using next generation sequencing.PLOS ONE / ed. Moses A.M. 2018;13(12):e0201270. doi: 10.1371/journal.pone.0201270
- Elmore J.M., Velásquez-Zapata V., Wise R.P. Next-Generation Yeast Two-Hybrid Screening to Discover Protein–Protein Interactions. Protein-Protein Interactions / ed. Mukhtar S. New York, NY: Springer US. 2023;2690:205–222. doi: 10.1007/978-1-0716-3327-4_19
- Kawai S., Hashimoto W., Murata K. Transformation of Saccharomyces cerevisiae and other fungi: Methods and possible underlying mechanism. Bioeng. Bugs. 2010;1(6):395–403. doi: 10.4161/bbug.1.6.13257
- Chen P. et al. Visualized investigation of yeast transformation induced with Li+ and polyethylene glycol. Talanta. 2008;77(1):262–268. doi: 10.1016/j.talanta.2008.06.018
- Yamakawa M., Hishinuma F., Gunge N. Intact Cell Transformation of Saccharomyces cerevisiae by Polyethylene Glycol. Agric. Biol. Chem. 1985;49(3):869–871. doi: 10.1080/00021369.1985.10866817
- Gietz R.D. et al. Studies on the transformation of intact yeast cells by the LiAc/SS-DNA/PEG procedure. Yeast. 1995;11(4):355–360. doi: 10.1002/yea.320110408
- Schiestl R.H., Gietz R.D. High efficiency transformation of intact yeast cells using single stranded nucleic acids as a carrier. Curr. Genet. 1989;16(5–6):339–346. doi: 10.1007/BF00340712
- Pham T.A., Kawai S., Murata K. Visualization of the synergistic effect of lithium acetate and single-stranded carrier DNA on Saccharomyces cerevisiae transformation. Curr. Genet. 2011;57(4):233–239. doi: 10.1007/s00294-011-0341-7
- Hayama Y. et al. Extremely Simple, Rapid, and Highly Efficient Transformation Method for the Yeast Saccharomyces cerevisiae Using Glutathione and Early Log Phase Cells. J. Biosci. Bioeng. 2002;94(2):166–171. doi: 10.1263/jbb.94.166
- Gietz R.D., Schiestl R.H. Quick and easy yeast transformation using the LiAc/SS carrier DNA/PEG method. Nat. Protoc. 2007;2(1):35–37. doi: 10.1038/nprot.2007.14
- Gietz R.D., Schiestl R.H. Large-scale high-efficiency yeast transformation using the LiAc/SS carrier DNA/PEG method. Nat. Protoc. 2007;2(1):38–41. doi: 10.1038/nprot.2007.15
- Gietz R.D., Schiestl R.H. High-efficiency yeast transformation using the LiAc/SS carrier DNA/PEG method. Nat. Protoc. 2007;2(1):31–34. doi: 10.1038/nprot.2007.13
- James P., Halladay J., Craig E.A. Genomic Libraries and a Host Strain Designed for Highly Efficient Two-Hybrid Selection in Yeast. Genetics. 1996;144(4):1425–1436. doi: 10.1093/genetics/144.4.1425
- Schneider C.A., Rasband W.S., Eliceiri K.W. NIH Image to ImageJ: 25 years of image analysis. Nat. Methods. 2012;9(7):671–675. doi: 10.1038/nmeth.2089
- Wickham H. ggplot2: Elegant Graphics for Data Analysis. Springer-Verlag New York, 2016.
- Singh, M.V., Weil, P.A. A method for plasmid purification directly from yeast. Anal. Biochem. 2002;307(1):13-7. doi: 10.1016/S0003-2697(02)00018-0
- Kreplak, J et al. A reference genome for pea provides insight into legume genome evolution. Nat. Genet. 2019;51:1411–1422. doi: 10.1038/s41588-019-0480-1
- Gietz R.D., Schiestl R.H. Applications of high efficiency lithium acetate transformation of intact yeast cells using single‐stranded nucleic acids as carrier. Yeast. 1991;7(3):253–263. doi: 10.1002/yea.320070307
- Dolgikh A.V., Dolgikh E.A. Searching for regulators that interact with BELL1 transcription factor and control the legume-rhizobial symbiosis development. Ecol. Genet. 2021;19(1):37–45. doi: 10.17816/ecogen51489
- Tripp J.D. et al. Enhancement of plasmid DNA transformation efficiencies in early stationary-phase yeast cell cultures: Enhancement of DNA transformation in yeast cells. Yeast. 2013;30(5):191–200. doi: 10.1002/yea.2951
- Ito H. et al. Transformation of intact yeast cells treated with alkali cations. J. Bacteriol. 1983;153(1):163–168. doi: 10.1128/jb.153.1.163-168.1983
- Klebe R.J. et al. A general method for polyethylene-glycol-induced genetic transformation of bacteria and yeast. Gene. 1983;25(2–3):333–341. doi: 10.1016/0378-1119(83)90238-X
- Németh T. et al. Enhancing the chemical transformation of Candida parapsilosis. Virulence. 2021;12(1):937–950. doi: 10.1080/21505594.2021.1893008
- Chan V. et al. The Effect of Increasing Plasmid Size on Transformation Efficiency in Escherichia coli. J Exp Microbiol Immunol. 2002;2:207–223
- Szostková M., Horáková D. The effect of plasmid DNA sizes and other factors on electrotransformation of Escherichia coli JM109. Bioelectrochem. Bioenerg. 1998;47(2):319–323. doi: 10.1016/S0302-4598(98)00203-7
- Bonett D.G., Price R.M. Confidence Intervals for Ratios of Means and Medians. J. Educ. Behav. Stat. 2020;45(6):750–770. doi: 10.3102/1076998620934125
- Yu S.-C. et al. Nutrient supplements boost yeast transformation efficiency. Sci. Rep. 2016;6(1):35738. doi: 10.1038/srep35738
- Causier B., Davies B. Analysing protein-protein interactions with the yeast two-hybrid system. Plant Mol. Biol. 2002;50(6):855–870. doi: 10.1023/A:1021214007897