Genomics Exam 1

Front 1

Basic goals of genome project

Back 1

1. Sequenced DNA that comprises major genome units
2. Annotate genome features
3. Analyze freq and distribution of features, make biological and evolutionary comparisons, and publish
4. Make sequence and annotations publicly available

Front 2

When did the HGP begin and end?

Back 2

1990 to 2003 (2 yrs ahead of schedule)

Front 3

Who headed the governments HGP?

Back 3

US dept of Energy and the NIH. PI was Francis Collins.

Front 4

What were the goals of the HGP?

Back 4

>identify all the approximately 20,000-25,000 genes in human DNA,
>determine the sequences of the 3 billion chemical base pairs that make up human DNA,
>store this information in databases,
>improve tools for data analysis,
>transfer related technologies to the private sector, and
>address the ethical, legal, and social issues (ELSI) that may arise from the project.

Front 5

What's a genome consortium?

Back 5

Association of scientists and public and private institutes working together to sequence and understand a genome for public release

Front 6

How do we use bioinformatics?

Back 6

• Store/retrieve biological information (databases)
• Retrieve/compare gene sequences
• Predict function of unknown genes/proteins
• Search for previously known functions of a gene
• Compare data with other researchers
• Compile/distribute data for other researchers
• Generate hypotheses for testing in the lab

Front 7

What does BLAST stand for?

Back 7

Basic local alignment search tool

Front 8

What can linear sequence of letters in the genome reveal about vying organisms?

Back 8

1. How info is carried out or expressed over time and space
2. How info is transformed over generations
3. Sequence alone is not the key to life

Front 9

What distinguishes genome level studies from genetic studies?

Back 9

Emergent properties of genome:
• Gene order along chromosome, gene frequency
• Genome characteristics: GC content, codon usage, transposons, can test horizontal gene transfer, changes in genome size
Scale/ Speed:
• GWAS – Genome Wide Association studies to find all genes associated with specific phenotype
• Complex genetic disease due to many interacting genes, many unknown genes, can quickly identify candidate genes
• Development of many genetic markers at once
• Find all differentially expressed genes related to alternative
phenotypic state

Front 10

Genomic information is differentially expressed over time and space in an individual. This reflects...

Back 10

Biological processes!:
>development (growth and differentiation)
>disease
>responses to environmental stimuli (think phenotypic plasticity)09

Front 11

ENCODE project determined

Back 11

That over 80% of the human genome's components have been assigned at least one biological function

Front 12

What's the take home message with gene regulation?

Back 12

Quite complex
Involves many interacting proteins

Front 13

Examples of differing expression in response to life stage or environmental stimuli

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1. Fetal vs adult regulation of gloving gene region
2. Daphnia pulex develops curved helmet and elongated tail spine in presence of predator chemical cues (34/107 genes were novel)
3. Frog genome response to Db infection - immune response not there at early stages. At later stages, differential gene expression more down regulated than up. Fungus winning! Gene suppression :(

Front 14

Epigenome is

Back 14

Modifications to genome fetching gene regulation without changing nucleotides
>involves DNA methylation or histone modification
>Methylation silences genes at CpG sites

Front 15

Genomic information is transformed over generations! This reflects processes of...

Back 15

1. Mutation
2. Population genetics
>Results in differences among populations and species

Front 16

Sanger sequencing

Back 16

Dideoxy chain termination sequencing

Front 17

ddNTP stands for ___________, and morphologically a ddNTP is ...

Back 17

Dideoxyribonucleotide
Missing 3' -OH group and terminates the DNA chain

Front 18

How was the HGP conducted?

Back 18

Hierarchical shotgun sequencing. Explain.

Front 19

Genomic library

Back 19

Contains entire genome in overlapping segments
>vector determines size of DNA insert that can be carried (100-200kb fragments into BAC vector during HGP)

Front 20

HGSP sequencing strategy

Back 20

Don't sequence everything
1. Take overlapping BACs and draw out tiling set (29k clones as opposed to 354k)
2. Shotgun sequence and computer assembly
3. More shotgun, gap closure, finishing

Front 21

Mapping:
Genetic maps vs physical maps

Back 21

Genetic:
>relative order of genetic markers
>Distance expressed in units of recombination frequency
>units = cM
Physical:
>Distances based on length of DNA sequence between genetic elements
>Units = kb, Mb, etc.

Front 22

Why does recombination help make genetic maps?

Back 22

1. Linked genes (alleles) transmitted as a single unit
2. Coinheritance implies two alleles on same chromosome

Front 23

Degree of crossing over (recombination) between two genes on the same chromosome is ...

Back 23

proportional to the distance between them. Map units, aka ____________, are ...

Hint 23

Centimorgans, 1 % recombination between 2 genes on a chromosome

Front 24

STSs

Back 24

Sequence tagged sites; known fragments of DNA of known chromosomal location, used to map BAC inserts

Front 25

HGP mapped BAC inserts by

Back 25

1. Using STSs
2. Digesting library clones with HindIIIo get DNA fragment fingerprint

Front 26

What is FISH and what is it used for?

Back 26

Fluorescent in situ hybridization; used to map BAC clones with no STSs

Front 27

What is a tiling set?

Back 27

Fragments with Minimal amount of overlap to allow seqs to be stitched back together

Front 28

Library

Back 28

~2kb fragment BAC inserts; ligate into a vector and sequence from either end (phage vector sequence is known)
>get about 800bp from each end, so close to 2000 bp total

Front 29

Contigs

Back 29

Computer program orders seqs based on overlap to form longest contiguous sequence

Front 30

Scaffold

Back 30

Overlapping clone sequences eliminate gaps, yield complete seq.; scaffold = merged contigs

Front 31

Define coverage

Back 31

Average number of times a base was sequenced

Front 32

Name some of the Bermuda Rules and who they applied to

Back 32

1996
-assembled seq greater than 1000bp deposited into public database every 24hrs
-no patents may be filed
-joint commitment of sequencing centers and funding agencies

Front 33

What did Celera do differently from the HGP?

Back 33

Whole genome shotgun sequencing instead of heierarchical shotgun sequencing; broke whole genome into pieces of different sizes, skipped mapping, Advanced computing assembled pieces into correct order

Front 34

Differences in next-gen sequencing?

Back 34

-Numerous technologies (454, Illumina, SOLID)
-much shorter fragments and longer runs
-thousands to millions of short frags done at once rather than 96 800bp reads
-real challenge is assembling these very short reads (75 to 400bp)

Front 35

Human genome is ~___ Gb but only ~__% codes for proteins;
Genome contains about how many genes?

Back 35

3.2; 1.5
25,000

Front 36

How do we find genes in a DNA sequence?

Back 36

Look for the DNA signatures of genes:
– Start codons
– Stop codons
– “open” sequence in between (ORF)
• These regions are potential genes
– i.e. they could produce proteins if proper promoters and translation initiation sequences are present.

• How many ways can you translate a DNA sequence?

Hint 36

6

Front 37

Ab initio methods of gene detection rely on...

Back 37

Signal and content:
Signal is presence of specific sequence (e.g., Prinbow box or -35
element, polyadenylation signal)
Content is statistical properties of protein-coding sequence itself (GC content, ORF, codon frequency, etc.)

Front 38

TATA box

Back 38

a DNA sequence (cis-regulatory element) found in the promoter region of genes in archaea and eukaryotes;[2] approximately 24% of human genes contain a TATA box within the core promoter

Front 39

Why is an initio gene prediction in eukaryotes more difficult than in prokaryotes?

Back 39

Less gene-rich
Introns
Complex promoter sequences

Front 40

What is an EST?

Back 40

Expressed sequence tag; partial sequence of a cDNA

Front 41

What are the difficulties associated with biological evidence?

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• Part of different cDNAs show similarity to same region of chromosome (alternative splicing).
• BLAST doesn’t distinguish splice sites, so intron boundaries often incorrect
• More often than not, have ESTs first, are often incomplete, 5’ end missing, so gene boundaries may be incorrect
• Predictions less accurate as move away from species

Front 42

Transposons

Back 42

Move within the genome and can insert themselves into various positions within and between chromosomes;
Usually exist in many similar copies within a genome ("families");
LINES and SINES are major families in humans

Front 43

Describe class I transposons

Back 43

Retrotransposons – resemble RNA viruses and move via an RNA intermediate (copy and paste)
•Usestheenzymereversetranscriptase(mayormay not encode RT).
•Mayormaynothavelongterminalrepeats(LTR’s)

Front 44

Describe class II transposons

Back 44

DNA transposons – move from one part of the genome to another by a cut-and-paste or copy and paste mechanisms
•Use the enzyme transposase thatallowsthemtojump (may or may not encode transposase)
• Cause small duplication at target site

Front 45

Retro transposons are transcribed by _____, the RT turns into _____ and gets ....

Back 45

Retrotransposons are transcribed by RNA pol II, RT turns into cDNA, reinserted into genome

Front 46

DNA Transposons move by two methods:

Back 46

nonreplicative (cut and paste) and replicative (copy and paste) transposition

Front 47

How are transposons identified in a new genome?

Back 47

Discovery: (e.g. homology based method) BLAST genome using transposon-specific sequences as queries, e.g., sequence for transposase, reverse transcriptase, integrase; scan surrounding regions for flanking repeats to iden3fy boundaries; (e.g. structure based method) look for long terminal repeats
Detection: (e.g., RepeatMasker/CENSOR) Iden3fy interspersed repeat sequences in genome that match consensus of transponsable element consensus sequence in RepBase databases