Anthony J. Wavrin Week 3: Difference between revisions
From OpenWetWare
Jump to navigationJump to search
(Added all of physiological factors) |
(Added all of RNA expression) |
||
| Line 5: | Line 5: | ||
*This study tests explores if increasing concentration of ammonia while keeping a constant influx will cause nitrogen related responses. | *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. | *The concentrations used resulted in testing from nitrogen limitation to nitrogen excess. | ||
*Overall, they conduct effects of physiological parameters, RNA expression, and enzyme | *Overall, they conduct effects of physiological parameters, RNA expression, and enzyme activites. | ||
==Physiological parameters== | ==Physiological parameters== | ||
| Line 11: | Line 11: | ||
**It is interesting to note that at the concentration of 61mM of ammonia, glucose becomes the limiting nutrient. | **It is interesting to note that at the concentration of 61mM of ammonia, glucose becomes the limiting nutrient. | ||
===Figure 1=== | ===Figure 1=== | ||
====A | ====A==== | ||
*The X-axis represents the increase in concentration of the ammonia. | *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 left represents the residual ammonia concentration. | ||
*The Y-axis on the right represents the biomass (dry weight). | *The Y-axis on the right represents the biomass (dry weight). | ||
| Line 19: | Line 19: | ||
*The residual ammonia concentration sky rockets after 61mM which is expected due to nitrogen excess. | *The residual ammonia concentration sky rockets after 61mM which is expected due to nitrogen excess. | ||
====B==== | ====B==== | ||
*The X-axis represents the increase in concentration of the ammonia. | *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 left represents the CO2 production. | ||
*The Y-axis on the far left represents the O2 usage. | *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. | *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. | *Concentrations above 44mM of ammonia have a relatively flat respiratory quotient, indicating carbon is the limiting nutrient. | ||
====C==== | ====C==== | ||
=====Left | =====Left Panel===== | ||
*The X-axis represents the increase in concentration of the ammonia. | *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. | *The Y-axis on the left represents the concentration of α-ketogluterate present. | ||
*As the ammonia concentration increases, α-ketogluterate concentration decreases until 61mM. | *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 | =====Middle Panel===== | ||
*The X-axis represents the increase in concentration of the ammonia. | *The X-axis represents the increase in the concentration of the ammonia. | ||
*The Y-axis on the left represents the concentration of glutamate present. | *The Y-axis on the left represents the concentration of glutamate present. | ||
*As the ammonia concentration increases, glutamate concentration increases until 61mM. | *As the ammonia concentration increases, glutamate concentration increases until 61mM. | ||
=====Right | =====Right Panel===== | ||
*The X-axis represents the increase in concentration of the ammonia. | *The X-axis represents the increase in the concentration of the ammonia. | ||
*The Y-axis on the left represents the concentration of glutamine present. | *The Y-axis on the left represents the concentration of glutamine present. | ||
*As the ammonia concentration increases, glutamine concentration increases continually. | *As the ammonia concentration increases, glutamine concentration increases continually. | ||
==RNA Expression== | ==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 <i>Xho</i>1-<i>Bam</i>1 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== | |||
Revision as of 21:25, 30 January 2013
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 conduct 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.
