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Author:Wallenius, Janne
Title:Lactobacillus rhamnosus GG:n geeniekspressiovasteet kasvuympäristön pH:n vaihteluun
Gene expression responses of Lactobacillus rhamnosus GG to pH changes of the growth environment
Publication type:Master's thesis
Publication year:2010
Pages:viii + 72 s. + liitt. 35      Language:   fin
Department/School:Elektroniikan, tietoliikenteen ja automaation tiedekunta
Main subject:Bioprosessitekniikka   (Kem-70)
Supervisor:Leisola, Matti
Instructor:Eerikäinen, Tero ; Pitkänen, Juha-Pekka
Electronic version URL: http://urn.fi/URN:NBN:fi:aalto-201203131500
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Location:P1 Ark Aalto  1547   | Archive
Keywords:Gene expression
geeniekpsressio
pH
Lactobacillus rhamnosus GG
qPCR
TRAC
atpA
aldB
cfa
groEL
hrcA
pstS
ldhD
Abstract (eng): Monitoring organism's gene expression can provide useful information on it's physiology in different phases of cultivation.
In this work expression of pH stress related genes found from the literature (atpA, aldB, cfa, groEL, hrcA and pstS) were studied as a relative gene expression study using ldhD as a reference gene.
Expression measurements were carried out using qPCR and TRAC methods.
Study organism was Lactobacillus rhamnosus GG, a renowned probiotic.
Base gradients caused by pH control in an industrial process and temperature gradients were simulated with a scale-down method.
A pH gradient was created in a plug flow reactor.
Furthermore, differences between laboratory cultivation medium and industrial cultivation medium were studied with batch cultivations.
In these cultivations the set point of pH control was oscillated.

In the scale-down experiments the responses of groEL, hrcA and atpA genes to temperature and pH changes were studied.
The expression of pstS gene was induced almost linearly in the chemostat cultivation experiments when base gradient in plug flow reactor was increased.
There was rather low correlations between the expressions of studied genes and surviving from acid stress test or freeze stability test, which were carried out from the samples of the chemostat cultivation experiments.
Based on gene expression measurements from the batch cultivations it seems that the type of the cultivation medium affects expressions of the studied genes.
In case of the industrial cultivation medium relative changes in gene expressions were less between different growth phases but expressions related to the reference gene were mainly higher compared to the laboratory medium.
Abstract (fin): Monitoroimalla organismin geeniekspressiota voidaan saada hyödyllistä informaatiota fysiologiasta bioprosessikasvatuksen eri vaiheissa.
Tässä työssä on tutkittu kirjallisuudessa aiemmin mainittujen pH-stressiin liittyvien geenien atpA, aldB, cfa, groEL, hrcA ja pstS ekspressoitumista suhteellisen geeniekspression menetelmällä käyttäen vertailugeeninä ldhD:tä.
Ekspressiomittaukset tehtiin käyttäen qPCR- ja TRAC-menetelmää.
Tutkittuna organismina oli probioottina tunnettu Lactobacillus rhamnosus GG (LGG).
Teollisen kasvatuksen pH:n säädössä esiintyviä emäsgradientteja sekä lämpötilagradientteja simuloitiin scale-down-menetelmällä.
Lisäksi tutkittiin laboratorioravintoalustan ja teollisen ravintoalustan välisiä eroja panoskasvatuksilla.

Scale-down-kokeissa havaittiin geenien groEL, hrcA ja atpA antavan vasteita lämpötilan ja pH:n muutoksiin. pstS-geenin ekspressio kasvoi lähes lineaarisesti kemostaattikokeissa tulppavirtausreaktoriin aiheutetun emäsgradientin kasvaessa.
Työssä tutkittujen geenien ekspressioiden yksittäiset korrelaatiot kemostaattikokeiden näytteille tehtyjen happostressi- ja pakastussäilyvyyksien välillä olivat vähäisiä.
Panoskasvatuksista tehtyjen geeniekspressiomittausten perusteella erilaisilla kasvualustoilla näyttäisi olevan vaikutusta tutkittujen geenien ekspressioon.
Teollisella alustalla ekspressioiden suhteelliset muutokset eri kasvuvaiheiden välillä olivat pienempiä, mutta vertailugeeniin verratut ekspressiot olivat pääsääntöisesti suurempia kuin laboratorioalustalla.
ED:2010-08-19
INSSI record number: 40188
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