Saccharomyces cerevisiae ve Metschnikowia pulcherrima İçeren Sıralı Fermantasyonda Karışık Başlangıç Kültürünün Kullanılmasına Yönelik Transkripsiyonel Yanıt

Abstract

Yeast has played a role in the production of basic foods for mankind throughout history. With the development of science and technology, the knowledge about yeasts has increased day by day. Today, problems encountered in yeast fermentation can be solved by genome-based studies, correct yeast selection and use. One of the current problems faced in the fermentation industry is the high alcohol level in the final product due to global warming and changing environmental conditions. In addition, the fermentation industry seeks to make a difference in the final product in the national and international market with minimum changes such as combinations and strain changes in the starter culture. Therefore, the use of non-Saccharomyces yeasts in fermentation for high quality wine production has become more common in recent years. Many studies have shown that complex yeast-yeast interactions occur when Saccharomyces cerevisiae and non-Saccharomyces yeasts are used as mixed cultures, resulting in significant differences in the metabolism of Saccharomyces cerevisiae.

This study aims to investigate the interactions occurring in the sequential fermentation of Saccharomyces cerevisiae and Metschnikowia pulcherrima strains at the transcriptomic level. For this purpose, the study is to consist of two parts. The first part intends to understand the effects of heat stress on the expression profile of genes involved in aromatic compound synthesis in the Ehrlich pathway. With this purpose, the expression profile of the aromatic compound synthesis-related genes (Ehrlich pathway genes) in the local yeast strain against heat stress, is investigated for the first time within the scope of this thesis, using the open source microarray data. The second part of the study, focuses on monitoring the transcriptional response of selected ARO8, ARO9, ADH2, ADH5 genes involved in aromatic compound synthesis during alcohol fermentation using sequential mixed starter culture (Metschnikowia pulcherrima + Saccharomyces cerevisiae).

In the study, 16 Saccharomyces cerevisiae and 6 Metschnikowia pulcherrima strains isolated from different regions of Anatolia and identified in previous studies have been used in Hacettepe University FoodOmics Research Laboratory Culture Bank. Since it is similar to the wine fermentation medium, all resuscitation and fermentation experiments have been carried out in grape juice. ARO8, ARO9, ADH2, ADH5 genes to be used in the study have been selected according to open source microarray data. RNA isolation, Reverse Transcription and RT-qPCR applications of the Saccharomyces cerevisiae and Metschnikowia pulcherrima strains have been carried out. Alcohol dehydrogenase II and V (ADH2, ADH5) and aromatic aminotransferase I and II (ARO8, ARO9) gene expression levels of Saccharomyces cerevisiae and Metschnikowia pulcherrima strains have been determined. Then, the technological properties of all Saccharomyces cerevisiae and Metschnikowia pulcherrima strains selected according to these gene expression levels have been investigated. S. cerevisiae pure culture and sequential mixed culture containing Metschnikowia pulcherrima + Saccharomyces cerevisiae strains have been created from the strains selected according to their gene expression level and technological characteristics. In order to monitor the fermentation process at the transcriptome level, RNA isolation and Reverse Transcription-qPCR have been carried out for gene expression analyzes from the samples taken on the 1st, 3rd, 5th and 7th days of fermentation. During the fermentation, daily brix, alcohol and pH measurements have been made and the fermentation process has also been monitored biochemically.

As a result, the study, has found out that the use of Metschnikowia pulcherrima + Saccharomyces cerevisiae sequential mixed culture caused changes in the expression level of genes involved in aromatic compound synthesis. The study also shows that low alcohol end product yield is closely related to the use of sequential mixed culture Metschnikowia pulcherrima + Saccharomyces cerevisiae. It has been determined that the average amount of alcohol measured during fermentation was reduced by about 30 percent with the use of sequential mixed cultures. The combination of one Saccharomyces cerevisiae and one Metschnikowia pulcherrima strain isolated and identified from original/local sources as a result of the study performed here is recommended as a starter culture combination for use in the fermentation industry.