Microbial Genomics Flashcards
What is a “genome?”
What is genomics?
- Genome
- entire complement of genetic information
- Includes genes, regulatory sequences, and noncoding segments of the genome
- Genomics
- Discipline of mapping, sequencing, analyzing, and comparing genomes
What are the 3 steps to sequencing a genome?
- Sequence the DNA
- Assemble the sequences into chromosomes or large gragments called “contigs”
- Predict genes (annotation)
What was the first “major” way to sequence a genome?
- Sanger Sequencing
- Used a Dideoxy analog to the dNTP that was missing an OH on the 3’ Carbon
- This made whateversequence stop at that analog
- Used a Dideoxy analog to the dNTP that was missing an OH on the 3’ Carbon
What is “sequencing” in Genomics?
Determining the precise order of nucleotides in a DNA molecule
How many templates should a Sanger-esque sequencing technique have?
1, purified template
What are some of the New sequencing techniques?
- 454 - 1 million reads/run; 700 bp length (no longer supported)
- Illumina - 4 billion reads/run; 300 bp length
- PacBio - 125,000 reads/run; 2000 bp length
What is the “modern” standard for genome sequencing?
Illumina
What is the 4th generation sequencing Technology?
Basically, how does it work and what is its limitation?
- Nanopore sequencing
- Detects eletrical current as DNA is threaded through a protein pore
- Detects 3 bases at a time
- Uses a Helicase (maybe) to feed DNA through the pore.
- High error rate - 5-38.2%
How do you assemble a genome, after sequencing?
- Align all reads to generate longer sequences - contigs
- Gaps may still remain where no reads are found
- Orientation can be assumed if closely related genomes have been completed
- Must close the gap with PCR and sequencing of PCR amplicons by Sanger Seq
How do you fill gaps in your genome after you’ve sequenced it?
- You could try to orient it if closely related genomes have been completed
- Have to clsoe with PCR and sequencing of PCR amplicons by Sanger seq
What is the last step in sequencing a genoeme?
Annotating (Bioinformatics)
What is Annotation?
What do the majoirty of genes encode?
What are functional RNAs?
- Converting raw sequence data into a list of genes or elements present in the genome
- proteins - mRNAs
- Functional RNAs = rRNAs, tRNAs, ncRNAs
What is one way we could use organisms with a similar ORF for sequencing?
- Most genes code for proteins, and functional Rnas like rRNAs and tRNAs are very conserved
- So, if organisms have similar ORFs you can could assume function of the ORF or even assume sequence…
What is synteny?
arrangement of genes in a genome
What are things you can do with a genome?
- Compare gene sequences between bacteria
- Compare mutations between strains
- compare arrangement of genes in genome
- Design new therapeutics to treat
How many bases are in a kilobase?
How many bases are in a megabase?
- Kilo - 1,000
- Mega - 1,000,000
The bigger your genome –> the [] number of protein-coding genes…
Increases
The larger you genome –> The [] % of gene-coding
smaller
What may be some reasons for why bacteria have such a low % of non-coding genes?
- They are small. Don’t want a big genome.
- The metabolic burden would be too great to have too large a genome.
What types of genes are typically the most abundant known class?
metabolic
Based on the standard deviations, there is basically no difference between the gene distribution in [] and [] …..
Bacteria and Archaea
What is the metagenome?
The total content of the organisms present in an environment
What is used in the bacterial world’s metagenomics to assign species?
16s rRNA
What can/can’t Metagenomics do?
- Metagenomics Can
- Sequence of DNA in a sample
- Diversity information
- Relative quantities of community members
- Metagenomics Can’t
- Tell you the useful members of a community
- Who is alive/who is dead
- What pathway are being used in an environement
- so use TRANSCRIPTOMICS instead!
