11: Gene Expression
- Page ID
- 24824
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- 11.1: Introduction
- DNA was described as a molecule consisting of 2 anti-parallel strands in a double helix by Francis Crick and James Watson. The elegant model illustrated the intrinsic redundancy that made DNA a suitable data storage vessel for genetic information. Francis Crick later posited a notion of how this information went from storage to an actual program that runs cells. Crick first posited it as a “sequence hypothesis”. This idea of information flow is called the Central Dogma of Molecular Biology.
- 11.2: Prokaryotic Transcription
- Glucose is the preferred energy source of cells. François Jacob and Jacques Monod sought to understand how bacteria made decisions to switch between different sugars as sources of energy. Jacob and Monod found that if glucose and lactose were presented as food for bacteria, there would be a biphasic growth pattern. Monod found that when lactose was the sole sugar, the expression of the enzyme β-galactosidase was induced and displayed a monophasic growth with a delay.
- 11.3: RNA Miniprep (Activity)
- RNA purification occurs similar to DNA preparations. A silica-based column is used where DNA is excluded from binding based on size and through an additional DNA digestion step using the enzyme DNase I. RNA is extremely fragile and prone to degradation. Thus, separate pipettes and plastics are usually used in labs to reduce the amount of exposure to environmental or experimental RNase. This page contains instructions on how to prepare RNA and perform a reverse transcription of eukaryotic mRNA
- 11.4: Quantitative Nucleic Acid Measurement
- Measurements can be made of individual genes of interest through PCR of those specific genes. A process known as Real-Time PCR or quantitative PCR (qPCR) is used to measure individual genes using fluorescence measurements. An intercalating agent that binds only to double-stranded DNA called Sybr Green is used in a qPCR machine that is measuring fluorescence after each cycle of PCR indirectly indicates the amount of amplified product.
- 11.5: Eukaryotic Transcription
- Unlike prokaryotic genes, the expression of genes in eukaryotic cells has complex systems of transcription factors that act on promoters to recruit RNA polymerases. Additionally, enhancer elements may reside many kilobases upstream of the promoter. These enhancers strengthen the transcription of the gene. In this case, transcription activator proteins or trans-activators augment the promoter activity.