14 (8) Genomes and genomics

Question 1

what is sangar sequencing

Answer 1

• Works almost same as PCR but uses dNTPs with fluorescent tags
  ○ Each base has their own colour
• Once we have all the sequences we run them on a gel
• Lazer excitation and detection by machine

Question 2

what is NGS (nest generation sequencing)

Answer 2

• We chop up the genome
• Sequence all the bits individually and put it back together
• Cell free (no cloning)
• High throughput of reads
  ○ Can sequence millions of fragments on a plate all together

Question 3

what are the steps to NGS

Answer 3

1) Break genome into small random, overlapping fragments
2) Sequence fragments
a. Each fragment Is called a “read”
3) Reassemble fragments
a. Using computer to identify overlapping sequences
b. Assemble reads together into a larger continuous sequence (contigs)
c. Contigs that are accurate and “complete” are “chromosome level”

Question 4

what is Illumina sequencing

Answer 4

• Much better than sangar
• Basically colour the dots ATCG accordingly

Question 5

Why do we still use sangar sequencing when next gen is around ?

Answer 5

• New gen more expensive for a single gene sequencing
• Sangar makes fewer errors per read
• Next gen is more time consuming

Question 6

what do we do with the sequences we get from these processes

Answer 6

• Study of entire genome
• Making function sense of entire genomes
• Bioinformatics
• comparative genomics

Question 7

what is Bioinformatics

Answer 7

• Annotating whole genome sequences
• Attempts to reveal relevant information abt the genomes
  ○ Protein coding gene
  ○ Non protein coding genes
  ○ Non-genic regions

Question 8

what is comparative genomics

Answer 8

• comparing genomes
• Shows how much genes have changed overtime between species
• Differences between genomes exposes a record of evolutionary history

Question 9

what are orthologs (orthologous genes)

Answer 9

○ Homo genes located in the same genetic locus in closely related species
- ex) hemoglobin protein found in humans and mice.

Question 10

what are paralogs

Answer 10

○ Homo genes duplicated and evolved to have diff functions
○ Arose from gene duplication in common ancester
- ex hemoglobin and myoglobin in humas

Question 11

orthologs and paralogs are subsets of what

Answer 11

homologs

Question 12

what are convergent genes

Answer 12

They look the same but don’t have the same ancestor (Opposite of homology)
- Eyes of an octopus and eyes of a human
- Wings of a bat and wings of a pigeon are convergent

Question 13

what are pseudogenes

Answer 13

Genes that don’t do anything
- Used to have functions in ancestors

Question 14

what is functional genomics

Answer 14

Study of interaction of gene producing –> interallelic affects
- Genes- where, when , how much (transcriptomics)
- Proteins- where, when, how quickly (proteomics)

Question 15

what is a contig

Answer 15

• The assembled piece of next generation sequencing
  
  • Collection of reads aligned together

Question 16

what are reads

Answer 16

• The pieces of DNA that get chopped up in a sequence

Question 17

what are traits of in situ hybridization

Answer 17

• Detects RNA in tissue
• Shows where genes are transcribed
• Design complimentary DNA or RNA that binds to our sequence
• Has labels that show us where the tissue is expressed (blue dye like)

Question 18

what is QPCR (quantitative PCR)

Answer 18

• Asks how much DNA/RNA was there in the first place (Relative measure)
• Every PCR cycle doubles the amnt of target DNA
• Duplicates DNA with an intercalating (sticks inside DNA) unit
• The more copies of DNA you have the quicker you’ll reach the detection threshold

Question 19

what are the uses of QPCR

Answer 19

• Detects small amnts of viral DNA
• detect transcription