Bch 447 lecture 6
Richard lenski in 1988 he decided to do an evolutionary experiment. So, he
started growing a culture of bacteria for many generations. He got 12 flasks
and placed e-coli b in there.
He seeded 12 flasks at 100ml each. The flasks had very low glucose...
Therefore limitting the growth. He also placed citrate in the medium to
chelate heavy metals as bacteria are sensitive to heavy metals.
He let flasks sit on a bench, and every day he would dilute the cultures
Every day there were 6.64 generations of e-coli. The experiments is still
going 60000 generations later.. It's 2014 now.
He would take alliquotes of each of the flasks and freeze them once every
The bacteria were under a lot of selective pressure... It makes sense that
the bacteria using the glucose more efficiently will grow more quickly and
After a year or two... All twelve of the cultures would become cloudier much
faster than at the beggining, probably using glucose more efficiently.
Most of the cultures had similar mutations.
Rbs mutation : ribose utilization operon... Important for ribose formation
SpoT mutation... It's a guanine nucleotide with two phosphate groups at the
5' and 3' end and it is an activator of operon transcription... It activates
genes for carbon metabolism.
MalT... Maltose... Important for the uptake of 6 carbon sugars. Mutations
here retrieve sugars other than glucose.... The bacteria fed on their dead
Also modifications of nad formation and pyruvate shuttling... Cell wall
biosynthesis genes. Plasmid regulation.. Topoisomerase helps unwind.
Basically all of the cultures had common mutations. But each flask had a few
mixtures of surviving bacteria with different genes.
One of the cultures after 30000 generations the ara3 type bacteria became
very dense. These guys had aqcuired a mutation allowing them to grow to
extensive densities. The culture was citrate plus.... The bacteria mutated to
use citrate as a carbon source along with glucose. Note that the other
cultures did not aqcuire this mutation.
Firstly, looking at the bacterial genomes they asked... What is the random
rate of mutation in these cultures... By comparing the genomes from previous
We know that mutations will occur.... And that some will be fixed. Further
more is there a background neutral mutation rate that is constant... Remember
this is different because there is hardly any junk DNA in e coli. It may be
that every mutation that occurs may modify a gene.
So, they decided to look at the synonymous mutations that occured in every
generation... Mutations that didn't change the amino acid sequence. So, what's the rate of fixation of these mutations.
So, they took all 941000 synonymous sites in e coli and compared them. About
1/5 of the genome.
They looked at 8 of the cultures. And they asked how many synonymous alleles
were there fixed accross all of them... They didn't use the other 4, because
four of the flasks had differences in their mutation rate.
So, they compared accross 300000 generations.
The equation is on the sheet... They found that the mutation rate was
consistent with the sequencing.
As per the equation if we are looking at neutral mutations the fixation rate
should be equal to the mutation rate. Which it is.
It appears that synonymous mutations here are essentially neutral.... Note
though that sometimes synonymous substitutions can be selected for.
Four of the cultures had mutations that changed the mutation rate 100 fold.
The mutation rate became 10^-8. It's likely because the bacteria wanted to
mutate faster Under the low glucose selective pressure. These e coli have way
more mutations per generation. The downside here is genetic load. The
prediction is that once the beneficial mutations required for survival in low
glucose arise... The population would revert the mutation rate to normal once
Wilga's said that we should see evidence in our 8 cultures that are now under
normal mutation rate.... We may be able to see evidence of mutate DNA
machinery... To show that perhaps those 8 flasks underwent a higher rate of
mutation and reached their improved phenotype... At which point they reverted
back to a normal mutation rate. Perhaps we can see a transient period of
hypermutation in the genome.
We don't select for mutations in DNA machinery.... But they accidentally
occur.... And we get hyper mutation.... If this hypermutation is
beneficial... Then that species survives.
Linkage disequilibrium... The bad mutation is carried along with the good
mutation.... That is the mutant DNA machinery inducing hypermutation get