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Microbiology > Lecture 10 > Flashcards

Lecture 10 Flashcards

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Biology 101 flashcards
1
Q

genome construction

A

“shotgun” approach
3 steps: fragmentation, sequencing, and assembly

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2
Q

fragmentation

A

cut the genome into small pieces
physical shearing and enzymatic methods

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3
Q

physical shearing

A

nebulization (DNA shreds to fit into a size-adjustable whole & most commonly used method)
sonication (soundwaves that fracture back bone of DNA, won’t lose any DNA)

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4
Q

Enzymatic methods

A

a chemical method
nuclease mixes (Fragmentase)
cuts DNA

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5
Q

DNA sequencing

A

determining nucleotide composition of DNA
first, second, and third generations

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6
Q

first generation sequencing

A

developed by Dr. Fred Sanger in 1977
PCR with Dideoxynucleotides (replaces OH with H– terminates DNA replication)
manual: four separate reactions one for each dideoxynucleotide & gel electrophoresis
automated: single reaction, uses fluorescent markers, and laser detection of sequence

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7
Q

second generation sequencing

A

Adv: massively parallel (different sequences run together) and high throughput (100 times faster & 33 times cheaper)
(Still) requires amplification of DNA samples
Methods: emulsion PCR and bridge PCR

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8
Q

bridge PCR

A

still use today
a flat piece of glass and bits of DNA attached to glass (called flowcell)
Illumina platform sequencing (get fluorescent signal where a nucleotide is added, read by computer, and rapid advancement)

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9
Q

third generation sequencing

A

adv: massively parallel (many sequences at the same time), extremely high throughput (“real-time” results), and does NOT require amplification of DNA samples (can sequence DNA immediately)
methods: SMRT technology (DNA polymerase) and nanopore technology (transmembrane proteins)

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10
Q

Assembly

A

reconstructing genomes
combine short, overlapping DNA sequences
computer aided sorting

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11
Q

Types of assembly

A

reference alignment
- comparison to known genome
- must be a closely related organism
De Novo assembly
- novel genome construction
- no closely relative required

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12
Q

De Novo assembly

A

how many times the genome is sequenced
- number of sequence reads per nucleotide
completion
- how much of the genome is sequenced
- closed (complete sequence, no gaps) v. drafted (incomplete sequence, gaps)
coverage
- how many times genome is sequenced (# of reads per nucleotide)

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13
Q

bioinformatics

A

analyzing and storing DNA/protein sequences
- utilizes powerful computational tools
comparative analyses
- genome size, content, and organization
bottleneck

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14
Q

Small genomes

A

140,000 to 1,000,000 bp
170 to 1,000 ORFs
- endosymbionts
- parasites

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15
Q

Large genomes

A

5,000,000 to 13,000,000 bp
5,000 to 9,000 ORFs
free-living organisms
- unpredictable environment

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16
Q

larger genomes

A
  • like in eukaryotes
    more genes
  • little “junk” DNA
  • DNA replication
    -translation (protein synthesis)
17
Q

gene content

A

annotation
- predicting functional genes from DNA sequence data
- based on comparative analysis
mystery genes
- 30% ORFs unknown roles
- e. coli

18
Q

genomics

A

all genetic information in the cell

19
Q

metagenomics

A

all genetic information in an environment
- pooled DNA from an environmental samples
- includes genes from many different organisms

20
Q

transcriptomics

A

expressed genetic information
- study of total gene expression (genomes = list of parts)
- Microarrays (gene chips)
applications: study of pathogenic bacteria and human cancer cells

21
Q

proteomics

A

translated genetic information
study of total protein production (includes protein structure, function, and regulation)
techniques: in vitro (separate and ID proteins) and in silico (predict proteins from DNA)

22
Q

Microarrays

A

silica chips containing different genes
sample mRNA hybridizes + fluoresces
- fluorescence indicates expression of each gene
- track expression levels of individual genes

23
Q

genome evolution

A

changes in genome content over time
through gene duplication and deletion
horizontal gene transfer

24
Q

gene duplication

A

segment of DNA copied in the genome
- main mechanism of new gene evolution
- one copy remains unchanged + functional
- one copy changes (mutates) to a new function

25
Q

gene deletion

A

loss of a segment of DNA in the genome
- common in endosymbionts and parasites
- dependence on hosts result in “useless” genes
– no selection pressure to retain these genes over time

26
Q

Pathogenicity islands

A

clustered virulence genes transmitted horizontally
pathogenic bacterial strains

27
Q

core genome

A

essential genes
in all strains of a species

28
Q

pan genome

A

non-essential genes
in some strains of a species

Microbiology (19 decks)

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