Lecture 6: Omics Approaches in Neuroscience

Question 1

What are major omics approaches?

Answer 1

genomics
transcriptomics
proteomics
metabolomics
epigenomics
miRNAomics

Question 2

What is the central dogma of molecular biology?

Answer 2

The central dogma of molecular biology is a theory stating that genetic information flows only in one direction: from DNA, to RNA, to protein, or RNA directly to protein

Question 3

What is genomics?

Answer 3

measuring of all the DNA

genome = complete set of DNA

this is the basis of our genetic material (very stable and easy to measure)

DNA is a double helix structure composed of 4 nucleotides

Question 4

What are some structures that regulate DNA transcription?

Answer 4

once the DNA is open specific structures regulate transcription

transcription factors gives the cell identity, cause all genes are not transcribed

enhancer elements bind to transcription factors near the promotor region (specific regions in specific cells that allow transcription to happen)

Question 5

What is the human genome project?

Answer 5

formally launched in 1990
international effort
completed 2003
cost $3 billion
now with modern costs can sequence a human genome for $600-1000

Question 6

What is the Sanger Sequencing method of DNA sequencing?

Answer 6

use primer elongation with chain termination

originally measured with gel electrophoresis

can measure now with flurochromes using capillary gel electrophoresis

Question 7

What is next generation sequencing?

Answer 7

can do in a day wat takes weeks in Sanger sequencing

passively parallel by measuring DNA fragments

require DNA to be fragmented into specific sized “reads”

during library preparation various sequences are added that are needed for the sequencing process (ex. primer sequences)

cutting things into fragments and measuring those fragments

Question 8

What are the steps of next generation sequencing?

Answer 8

1. sample pre-processing
2. library preparation
3. sequencing
4. bioinformatics

Question 9

Where to SNPs occur?

Answer 9

almost once in every 1,000 nucleotides

roughly 4 to 5 million SNPs in a person’s genome

to be classified as a SNP, a variant is found in at least 1 percent of the population

Question 10

What methods do “Ancestry” and “23 and Me” use to help measure DNA?

Answer 10

phenotyping

based on sampling, SNP analysis, not full genome sequencing

Question 11

What are the steps in genotyping and genome wide associations (GWAS)?

Answer 11

1. data collection
2. genotyping
3. quality control
4. imputation
5. association testing

Question 12

How is genotyping performed?

Answer 12

genotyping here done with microarray
much cheaper than sequencing

Question 13

What is P-value?

Answer 13

p-value is the probability of an event happening due to chance

Question 14

What is the definition of genomics and epigenomics?

Answer 14

the study of the DNA sequence and associated heritable biochemical modifications

Question 15

What is the definition of transcriptomics?

Answer 15

the study of the RNA molecules present in a sample

Question 16

What is mature mRNA?

Answer 16

mature mRNA has no introns

it is mature mRNA that is often measured in transcriptomics

Question 17

What is the y-axis and x-axis in transcriptomic data?

Answer 17

y-axis: each line is a differentially expressed gene

x-axis: each line is a biological replicate

Question 18

What is proteomics?

Answer 18

transcript measurement gives only a rough estimate of its level of translation into a protein

many proteins experience post translational modifications (ubiquitination, phosphorylation, sumolation, glycosylation, acetylation, methylation, nitrosolation)

many transcripts give rise to more than one protein (alternative splicing)

Question 19

What are ways of measuring proteins?

Answer 19

immunoassays
western blotting
ELISA
mass spectrometry

Question 20

What is Western blotting?

Answer 20

ways to measure one protein at a time

same kind of thing as PCR

Question 21

What is ELISA?

Answer 21

can measure powerful quantitive changes in proteins

very expensive and can only measure one thing at a time

Question 22

What is mass spectrometry?

Answer 22

by measuring mass of structure, can figure out amino acids that make up protein

Question 23

What are the steps of mass spectrometry?

Answer 23

1. digest protein into pieces
2. liquid chromatography coupled to electrospray ionization
3. measure peptide sequence with MS/MS
4. identify and quantify proteins
5. conduct bioinformatics

every modification has to be studied seperately

Question 24

What is proteomics used for?

Answer 24

often used to identify biomarker ex. blood serum protein linked to disease

can be used to find “protein of interest” for disease or biological process

can measure protein changes to understand how a cell, tissue, system is changing

Question 25

What are the steps of preparing tissues and cells to perform omics?

Answer 25

mouse –> dissect –> brain –> dissociate tissue –> single cell suspension –> FACS –> CNS cells –> conduct transcriptomics, proteomics, or other omics

Question 26

What is fluorescent activated cell sorting (FACS)?

Answer 26

sorting out cells of interest

using defector plates, cells induced with a positive charge go one way and cells induced with a negative charge go the other

Question 27

What is the single cell revolution?

Answer 27

bulk sequencing: blend of all colors at the same time, cannot identify cell population or how they change

single-cell sequencing: discover new cells by measuring individually