April 1

Question 1

What is the transcriptome?

Answer 1

It is all RNA molecules (mRNA, tRNA, rRNA, etc

Question 2

How do we do qRT-PCR?

Answer 2

This quantitative reverse transcriptase PCR has random primers binding to RNA, then reverse transcriptase will make cDNA. Then make second cDNA strand for PCR

Question 3

What is the most abundant RNA in bacteria?

Answer 3

Ribosomal (rRNA)

Question 4

As we have lots of ribosomal RNA, if try to make cDNA of mRNA, we don’t get much. How can we increase yeild of cDNA from mRNA?

Answer 4

We use not so random primers, so we first get random primers, and duplicate only the ones of interest, making a batch of primers highly composed of primers for mRNA

Question 5

What is RNA-seq concept?

Answer 5

1. Firstly get reads, and align them with the genome
2. We have + (sense) strands and - (antisense). We look at how many repeats at least overlap with the gene, intergenic sequence or whatever.
3. We can test many conditions, see which genes, non coding RNAs, or etc that are up or down regulated. Can also see if antisense is upregulated, as antisense strands will form dsRNA with sense strands, so they indirectly reduce protein made from sense mRNA strands