mRNA

Question 1

What can mRNA be helpful for studying ?

Answer 1

• Gene activity and expression
• Gene regulation
• Transcriptional activities
• Post-transcriptional modification
• Regulation of translation

Question 2

What are the two ways of extracting and isolating total RNA from a cell ?

Answer 2

• Without a kit (biochemically), using guanidinium thiocyanate, phenol and chloroform.
• With a kit (by affinity), using silica-based columns.

Question 3

What are the steps of silica based columns extraction of RNA ?

Answer 3

• Cells are lysed
• Proteinase K degrades proteins/enzymes
• RNA binds to the column
• Debris and non RNA elements are washed away
• RNA can be harvested by elution in water

Question 4

What are the 2 mRNA quantification technics ?

Answer 4

• Spectrometry
• Fluorimetry

Question 5

How does fluorimetry mRNA quantification work ?

Answer 5

• Fluorochromes bind specifically to RNA/DNA and emit fluorescence which is detected by the fluorometer
• Fluorescent signal proportional to RNA/DNA content

Question 6

What mRNA quality verification technic exists ?

Answer 6

Fragment analysis :
- RNA integrity analysis is crucial in the RNA : it shows how much RNA is intact and usable in the form of a number : RIN

Question 7

What is a PCR ?

Answer 7

A PCR (polymerase chain reaction) is a molecular biology technique to amplify and detect specific DNA and RNA sequences.

Question 8

What 2 types of PCR exist ?

Answer 8

• Qualitative PCR : Endpoint PCR
• Semi quantitatively using fluorescent dyes (SYBR) : qPCR and RT-qPCR : Real time PCR