Unit 2 - Learning Objectives

Why is the genome important for DNA storage?

It contains bases that are transcribed into RNA and translated into amino acid chains that make up the protein necessary for cellular structure, function, survival and reproduction

What can genetic sequencing tell us?

It can tell us about a microorganism’s characteristics such as identity, metabolic pathways, energy production, nutrition requirements, and phylogentic relationships without having to culture the cell

What can performing targeted sequencing allow us to do?

It allows us to compare one gene in many organisms and look for mutations with a gene in a population of organism

What type of genomic sequencing would be best to perform if you were looking to identify genetic differences between different biochemical pathways for the same process in different microorganisms

Targeted sequencing

What can performing whole genome sequencing allow us to do?

It allows us to study all genes in one or more organisms and to discover regulation, pathways, and metabolism in uncultured organisms

If you were trying to determine a characteristic of an organism, such as it’s metabolic pathways, which type of genetic sequencing would be best to use?

Whole genome sequencing

What are the features of an ORF (open reading frame)?

A ribosomal binding site followed by a start codon, which begins the coding sequence which is ended by the stop codon

How does the computer analyze DNA sequencing data to find the ORF?

A possible start codon is identified → a possible stop codon is identified → computer counts the number of bases between possible start and stop codons to ensure they are a multiple of 3 → identifies a possible ribosomal binding site → computer calculates codon bias in ORF → computer decides if ORF is likely to be genuine → computer adds it to the list of probable ORFs

Why can parasitic bacteria exist with smaller genomes than microbes that are free living?

Parasitic bacteria require fewer metabolic pathways than free living bacteria because they can synthesize more nutrients directly from the host

What can genome sequences tell us about uncultured bacteria?

Their identity, metabolic pathways, energy production, nutrient requirements, and phylogenetic relationships

Why can the vast majority of microbes only be studied using genetic sequencing?

99% of all prokaryotes cannot be cultured in the laboratory because they live in complex communities that cannot be replicated in cultures

What does branch length tell us about the evolutionary relationship between organisms?

It is proportional to the evolutionary distance of the organism at the end of the branch to the organism present at the node

What information does comparative genomics provide us when comparing two organisms?

It allows us to identify differences between two organism’s genomes which can help us determine the effect of different genes and how different organisms relate to each other

Which gene is usually used to determine evolutionary relatedness between organisms?

Small subunit ribosomal RNA gene

Why is small subunit ribosomal RNA gene usually used for determining evolutionary relation between genes?

This gene has the same functions within all organism and can be shorted to just looking at variable regions to reduce needed computing power as the other sections rarely change

Why is using genes with the same function important for determining evolutionary relation between species?

Genes that have different functions usually evolve in different ways

What are other genes besides small subunit rRNA genes that are used for phylogentic comparison?

Genes for ATPase and glycolysis enzymes

What are advantages of having a large genome?

There are lots of genes that allow survival in a variety of habitats

What is a disadvantage of having a large genome?

Carrying a large genome requires more resources than carrying a small genome

How are microorganism’s identified through nucleic acid hydribization?

DNA from two organism’s, one known and one unknown, is added together in solution and heated so that DNA strands separate before slowly being cooled to reform them - matching strands for different organism will hybridized when cooled and if >70% of DNA from each organism hybridizes, it indicates they are the same species

How are microorganism’s identified through Southern blotting?

Flurescently tagged DNA are used to identify an unknown species based on if that specific sequence of DNA is present

How are microorganism’s identified through DNA chips?

A chip contains many different fluorescently tagged DNA probes that bond to species specific DNA sections of an unknown pathogen to identify the species

How are microorganism’s identified through ribotyping?

Looks at the rRNA of the small ribosome subunit to determine the phylogeic relationship between organisms and to use the specific sequence to determine the species

How are microorganism’s identified through flurescent in site hybridization (FISH)?

Cells are treated to allow DNA probes to bind with DNA to determine the identify, abundance,and relative activity of microorganisms within an environment

What are the steps of genome sequencing?

Sequences are aligned into “contigs” → scaffolds → “gap closure” and “finishing” → gene prediction → annotation

What happens during the “gap closure” stage of genome sequencing?

Regions of low sequence data are identified

What happens during the “finishing” stage of genome sequencing?

The scaffolds are synthesized into a draft genome

What happens during the “gene prediction” stage of genome sequencing?

Computer predicts the location of genes

What happens during the “annotation” stage of genome sequencing?

Function of genes are assigned based on sequence similarities between the sequenced genome and the unknown genome

True or false: alignments can only be done for sequences read from a single microorganism, not for an entire microbial community containing hundreds of species

False