Anthony J. Wavrin Week 3

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==Introduction==
==Introduction==
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This article is exploring one of the possible explanations of how nitrogen, used in the form of ammonia in this study, can effect <i>Saccharomyces cerevisiae</i>.  It is well known that nitrogen is an essential nutrient that can increase growth, as utilized in fertilizer.  It is hypothesized that the actual influx of nitrogen may cause growth instead of the concentration.  This study tests explores if increasing concentration of ammonia while keeping a constant influx will cause nitrogen related responses.  The concentrations used resulted in testing from nitrogen limitation to nitrogen excess.  Overall, they conduct effects of physiological parameters, RNA expression, and enzyme activites.
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This article is exploring one of the possible explanations of how nitrogen, used in the form of ammonia in this study, can effect <i>Saccharomyces cerevisiae</i>.  It is well known that nitrogen is an essential nutrient that can increase growth, as utilized in fertilizer.  It is hypothesized that the actual influx of nitrogen may cause growth instead of the concentration.  This study tests explores if increasing concentration of ammonia while keeping a constant influx will cause nitrogen related responses.  The concentrations used resulted in testing from nitrogen limitation to nitrogen excess.  Overall, they analyze effects of physiological parameters, RNA expression, and enzyme activites.
==Physiological parameters==
==Physiological parameters==

Revision as of 00:27, 31 January 2013

Contents

Introduction

This article is exploring one of the possible explanations of how nitrogen, used in the form of ammonia in this study, can effect Saccharomyces cerevisiae. It is well known that nitrogen is an essential nutrient that can increase growth, as utilized in fertilizer. It is hypothesized that the actual influx of nitrogen may cause growth instead of the concentration. This study tests explores if increasing concentration of ammonia while keeping a constant influx will cause nitrogen related responses. The concentrations used resulted in testing from nitrogen limitation to nitrogen excess. Overall, they analyze effects of physiological parameters, RNA expression, and enzyme activites.

Physiological parameters

  • The concentrations of ammonia used were 29, 44, 61, 66, 78, 90, 96, 114, and 118mM.
    • It is interesting to note that at the concentration of 61mM of ammonia, glucose becomes the limiting nutrient.

Figure 1

A

  • The X-axis represents the increase in the concentration of the ammonia.
  • The Y-axis on the left represents the residual ammonia concentration.
  • The Y-axis on the right represents the biomass (dry weight).
  • The Y-axis on the far right represents the ammonia flux, which is calculated using ammonia concentration, residual ammonia concentration, and biomass.
  • As ammonia increased to ammonia saturation, there was an increase in biomass, but stayed relatively constant after ammonia excess (>61mM).
  • The residual ammonia concentration sky rockets after 61mM which is expected due to nitrogen excess.

B

  • The X-axis represents the increase in the concentration of the ammonia.
  • The Y-axis on the left represents the CO2 production.
  • The Y-axis on the far left represents the O2 usage.
  • The Y-axis on the right represents the respiratory quotient, which is CO2 production/ O2 usage.
  • Concentrations above 44mM of ammonia have a relatively flat respiratory quotient, indicating carbon is the limiting nutrient.

C

Left Panel
  • The X-axis represents the increase in the concentration of the ammonia.
  • The Y-axis on the left represents the concentration of α-ketogluterate present.
  • As the ammonia concentration increases, α-ketogluterate concentration decreases until 61mM. This represents the conversion of α-ketogluterate with nitrogen to form glutamate and eventually glutamine.
Middle Panel
  • The X-axis represents the increase in the concentration of the ammonia.
  • The Y-axis on the left represents the concentration of glutamate present.
  • As the ammonia concentration increases, glutamate concentration increases until 61mM.
Right Panel
  • The X-axis represents the increase in the concentration of the ammonia.
  • The Y-axis on the left represents the concentration of glutamine present.
  • As the ammonia concentration increases, glutamine concentration increases continually.

RNA Expression

  • To measure the levels of RNA of nitrogen related genes, northern analysis was performed.
  • RNA was detected using labeled phosphate oligonucleotides or 2.5kB Xho1-Bam1 DNA fragments.
  • X-ray films were utilized to quantify the RNA Detected.

Figure 2

Left Panel
  • The X-axis represents the increase in the concentration of ammonia.
  • The Y-axis represents the % expression of a given RNA, using ACT1 and H2A-H2B as internal controls.
  • As the ammonia concentration increases past 61mM, GDH1 decreases while GDH2 increases, indicating that GDH1 is repressed by excess nitrogen while GDH2 is induced by excess nitrogen.
Middle Panel
  • The X-axis represents the increase in the concentration of ammonia.
  • The Y-axis represents the % expression of a given RNA, using ACT1 and H2A-H2B as internal controls.
  • Both GAP1 and PUT4 "peak" and 44mM and decrease til 78mM then become relatively stable, indicating that they are repressed by excess nitrogen.
Right Panel
  • The X-axis represents the increase in the concentration of ammonia.
  • The Y-axis represents the % expression of a given RNA, using ACT1 and H2A-H2B as internal controls.
  • GLN1, HIS4, and ILV5 all peak around 66mM and 78mM and stay at a high expression until the highest concentration (118mM), in which the expression decreases drastically.

Enzyme Activities

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