nm0238: okay good morning everyb-, [0.3] good af-, [0.8] it's Friday i don't 
even know whether it's morning or afternoon [0.5] good afternoon everyone [0.9] 
er i hope you're all [0.6] comfortable and have switched your mobiles off [1.3] 
so that we have a nice quiet time apart from me [1.1] er [0.6] first of all a 
reminder [1.0] about the practical sessions this afternoon and of course i'll 
put this up again later [0.9] er [2.0] group D are doing practical three in A-M-
S-G-eight [1.3] group A are over in Plant Sciences lab B doing [0.8] the er [0.
2] mitosis practical i think [1.8] the miosis practical and group C are doing 
the wash up practical one with me [0.5] in G-four [5.3] we're also er [0.2] we 
also welcome to this session [1.0] a member of the er Centre for Applied 
Language Studies staff [0.8] who thinks er that this [1.2] lecture may be worth 
recording [1.5] well there we go [4.6] my topic [0.3] for the first session 
this afternoon [0.8] is D-N-A and information [2.1] we've spent a long time now 
[0.4] er talking about genetics [1.5] about [0.7] m-, [0.2] genetics at the 
level of the [0.4] individual [0.3] at the level of the phenotype [0.7] at the 
level of the population [0.2] last week [1.3] [0.2] and at the level of the 
gene [0.8] undefined idealistic gene [0.5] Mendel's gene [2.2] which we've 
identified so far with a locus on a chromosome [1.8] but always at the back of 
our minds has been the fact that we all know that we live in the post-D-N-A age 
[1.0] and there is another way of explaing genetic behaviour [0.5] and that is 
to explain it at the molecular level [0.7] to explain it in terms of this 
molecule [0.5] deoxyribonucleic acid [1.5] D-N-A [2.1] and at about the same 
time that we discovered that D-N-A was the genetic informa-, the genetic er 
material [1.4] we also [0.6] were powerfully influenced by a theory [0.5] that 
said genetic material must consist of information [0.8] this was [0.4] a theory 
[0.8] which was based on [0.8] early computer theory [0.8] er [0.6] but was 
much supported by [1.3] the famous physicist Erwin Shrödinger [0.9] who during 
the last war [0.4] lived understandably away from Germany in Dublin [0.5] and 
he gave a very powerful [0.2] series of lectures in Dublin called What is Life 
[0.8] in which he propounded the 
view [0.7] that [0.3] at the bottom of life was genetics [0.4] and at the 
bottom of genetics had to be some sort of information [0.5] so we'll look at 
that information [0.3] paradigm or model [0.6] of er [0.6] genetics alongside 
[0.5] er [0.3] information about D-N-A [1.3] now i know that there are some of 
you [0.6] who will have been talking about D-N-A [0.5] for many years now and 
will be [0.4] if an-, if y-, if it's possible to be [0.6] and since i'm a D-N-A 
biochemist i don't think it's possible to be [0.8] er may actually be bored 
with it [0.7] well we'll try and [0.3] er liven it up as much as we can [0.2] 
when we talk [2.6] the substance of this lecture [1.0] has three parts [1.0] er 
we're going to remind ourselves what the evidence is [0.4] for D-N-A as the 
genetic material [0.9] then we're going to look at nucleic acids as molecules 
[1.1] and finally we're going to look at the functionality of D-N-A [1.0] how 
this model of D-N-A as information maps onto biochemical mechanisms [1.4] and 
in my second talk this afternoon we'll move on to talk a bit more about that [3.
6] so the evidence first of all then for D-N-A 
as the genetic material [3.3] and there are two [2.3] two canonical experiments 
[0.9] er [1.1] which [0.5] are described [0.2] in [1.1] chapter sixteen of 
Campbell [0.9] er and one of which we have already talked about so i will talk 
about the second today [1.1] the first one is the transformation experiment [0.
8] which we talked about when we were talking about transformation [0.5] in our 
consideration of bacterial genetics [0.6] remember transformation is when [0.3] 
an external D-N-A source [0.6] er [0.3] is added to a bacterium the bacterium 
takes up the D-N-A [0.4] and changes [0.2] its er [0.7] phenotype [0.3] in 
response to that new genetic information [1.2] and you'll remember [0.6] that 
this was originally [0.3] studied by the [0.2] English bacteriologist Griffith 
[0.9] when he looked at rough and smooth pneumococci [0.5] bacteria that caused 
[0.3] lung disease [0.5] and killed [0.2] mice [1.4] if they were smooth but 
not if they were rough [0.8] and then he showed that an extract of D-N-A from 
the smooth bacteria [0.4] could transform the rough bacteria [0.3] to make 
those equally pathogenic [1.3] now of course at that point you remember [0.3] 
he didn't 
know anything about what it was [0.4] that was the transforming agent as he 
called it [1.1] er but [0.3] subsequently [0.4] three researchers [0.2] in 
Baltimore [0.9] Avery MacLeod and McCarty [0.4] went through a series of 
experiments [0.3] where they digested [0.6] that [0.4] er preparation of 
transforming agent [0.5] with different enzymes [0.7] so they [0.2] digested it 
with [0.2] enzymes which [0.7] er broke down protein [0.4] which broke down R-N-
A [0.3] which broke down sugars [0.3] which broke down D-N-A [0.5] and they 
discovered that the only set of enzymes which killed [0.7] the ability of the 
extract to transform [0.4] were the [0.5] D-N-A [0.2] breaking down enzymea D-N-
A-ases [0.6] nucleases [1.2] and so they came to the conclusion that D-N-A [0.
3] must be [0.4] the transforming agent [0.5] and hence [0.3] what it was [0.7] 
that was in [0.9] the smooth bacteria [0.3] which [0.5] when donated to the 
rough bacteria made them pathogenic [2.4] the second [1.0] experiment [0.5] is 
known [0.3] throughout biochemistry as the blender experiment [1.4] because [0.
2] er [0.2] a material part in the experiment is taken by an ordinary kitchen 
blender [2.4] and that is described [1.1] on this overhead which you have a 
copy [0.4] of [1.3] in your handout [1.2] it's more properly known at the 
Hershey-Chase experiment [1.3] not for those of you who come from the other 
side of the water because it has anything to do with chocolate bars [0.5] but 
because [0.9] there was a man called Hershey and a man called Chase [0.3] in 
fact a woman called Chase [0.6] er who did this experiment [1.6] and this 
brings us back [0.5] again [0.5] to our consideration of bacterial genetics 
because it involves one of those organisms [0.4] we talked about in bacterial 
genetics [0.3] that is the bacteriophage [1.1] and you'll remember this picture 
[0.5] of a bacteriophage looking rather like [0.6] a lunar module [1.5] here 
attaching to a host bacterium [1.9] well what Hershey and Chase tried to do [0.
5] was to show what it was [0.9] that a phage [0.3] injected into [0.4] the 
host bacterium [0.6] which enabled that host bacterium to become then [0.4] a 
machine [0.3] for constructing new phage [1.8] and they did that using what was 
at that time very new technology [1.6] they [0.2] er [0.2] radioactively 
labelled [0.3] their phage [1.1] and they used [0.7] two radioactive labels [0.
9] thirty-five-sulphur [0.7] 
and [0.4] thirty-two- [0.2] phosphorus [1.1] so the [0.2] the upper line of 
this [0.3] describes what happens [0.4] er with thirty-five-sulphur [0.7] and 
the bottom line [0.4] describes what happens with thirty-two-phosphorus [0.8] 
the reason they used the two labels [0.6] is [0.5] that [0.4] there is [0.2] 
quite a lot of sulphur [0.5] in [0.9] proteins [0.3] coming from the amino 
acids methionine and cysteine [1.2] and very little phosphate [1.3] whereas 
there is a great deal of phosphate [0.6] in nucleic acids [0.3] and no sulphur 
[1.4] so using these two labels [0.6] the thirty-five-S showing what the 
protein is doing [0.5] and the thirty-two-P [0.3] showing what the [0.3] 
nucleic acid is doing [0.4] they hoped to be able to show what it was [0.3] 
that got into the bacterium [0.5] and caused [0.4] the er [2.2] caused the er 
[1.0] genetic changes in the bacterium [1.2] so [0.8] in both experiments what 
you do is to take your radioactively labelled phages [0.6] in a a flask you 
infect [0.2] some E-coli bacteria with those radioactively labelled phages [0.
4] and you allow them to attach [1.6] and even after they've injected their 
genetic information into the host cell [0.4] they will 
remain attached so we have to detach them [0.6] and the way we do that is to 
use the sheer forces generated by an ordinary kitchen blender [0.8] so you put 
your mixture [0.2] into [0.3] the blender [0.7] and you whirl it around a bit 
[0.9] and then you use a centrifuge [0.5] to separate [0.2] the supernatant [0.
4] which contains the phage particles that have broken off from the outside of 
the bacteria [0.5] and the pellet [0.2] which contains the bacteria [1.6] and 
when you do that [0.6] and you do it with thirty-five-sulphur [0.8] then all 
the [0.5] radioactivity remains in the supernatant [0.7] but when you do it 
with radioactive phosphorus [0.5] all the radioactivity [0.5] is in the pellet 
[0.8] so you know that it's the phosphorus [0.4] from the bacteria from the 
phage that's entered the bacteria [0.3] and not the thirty-five- [0.4] S [0.5] 
and so we conclude [0.3] that it's the nucleic acids in the phage [0.4] that 
are carrying the genetic information [0.2] and not the protein [1.3] okay [0.4] 
another very [0.2] simple experiment it looks t-, almost trivially simple to us 
now [0.6] er [0.5] but it was very instrumental [0.5] in showing [0.5] that D-N-
A 
was the genetic information so now we were able to show [0.6] that in bacterial 
transformation [0.2] and in phage infection [0.7] the genetic information that 
was being transferred [0.7] was [0.3] a nucleic acid [0.4] and in particular [0.
2] D-N-A [5.2] so we now know that D-N-A is the genetic material [0.7] what 
sort of stuff [0.5] is D-N-A [3.2] and in general what sort of [0.4] stuff [0.
3] are nucleic acids [2.4] well nucleic acids [0.5] as you all know are 
polymers [1.5] and in the case of D-N-A strikingly long [0.2] polymers [0.5] 
polymers being [0.2] chemicals which are constructed from repeating simpler 
units [4.9] the base unit [0.7] of a nucleic acid is what we call a 
polynucleotide [1.8] okay and so [0.3] this is an appropriate point to remind 
ourselves what a nucleotide is [11.1] 
and basically a nucleotide [1.3] is a combination of three things [0.5] a base 
[0.7] a sugar [0.6] and a phosphate [0.9] and for those of you who like like 
like their chemistry really simple and i'm sure that applies to a lot of you [1.
3] we can think [0.2] of a nucleotide [0.2] as being constructed in a very 
simple way [1.2] with a base [0.9] here [0.8] bonded [0.8] by a single bond [0.
7] to a sugar [5.2] and that sugar being again bonded [0.6] by a single bond [0.
2] to a phosphorus [1.9] okay [1.2] now this bond here [0.2] is an oxygen phos-,
[0.2] er carbon-oxygen-phosphorus bond [0.6] this bond here [0.4] is a carbon-
nitrogen bond [0.5] but er [0.3] that's for the chemists to worry about most of 
us [0.8] not too worried about that [2.7] but those of us who [0.6] w-, [0.7] 
and all of you should have at least some familiarity with what those chemicals 
look like [0.4] whether you like it or not [6.8] we'll look at [0.8] a slightly 
more complicated version [0.3] of what those chemicals look like [5.1] okay so 
[0.4] our bases [1.7] right [0.4] can be any of [0.9] in the case of D-N-A [0.
5] four [0.2] different bases [0.5] 
adenine [0.2] guanine cytosine [0.3] and [0.2] thymine [2.7] and in the case of 
R-N-A [0.5] any of the four bases adenine guanine cytosine or uracil [4.3] the 
difference between those [0.4] different bases [0.8] what we have there is a 
picture of thymine and uracil [2.1] but adenine [2.5] is a typical base [0.7] 
of the other kind so let's just draw [1.2] the two kinds of D-N-A bases you've 
seen thymine which looks like this [12.4] okay that's thymine [1.5] and thymine 
is what we call a pur-, [0.2] a pyrimidine base [3.6] and it just has a single 
[1.4] aromatic ring [1.5] okay [0.2] and that aromatic ring contains two 
nitrogens [0.9] the difference between thymine [0.4] cytosine and uracil [0.3] 
which are the three pyrimide bases [0.4] lies in what substituents we have [0.
6] in the case of thymine and uracil it's two oxygens [0.5] in the case [0.2] 
of er cytosine it's an oxygen and a nitrogen [2.2] if we look at the pyrimidine 
bases they're slightly different and slightly more complicated [17.6] just try 
and draw the bonds right [0.7] okay that's adenine [3.5] which is the simplest 
of these more complex bases which are 
called purines [2.2] okay [0.3] so there are two types of bases purines and 
pyrimidines [2.1] one is a single ring [0.7] and the slightly longer name [0.4] 
smaller structure longer name [0.5] and the other one has a double ring [0.5] 
and a shorter name [1.2] okay as well as thymine in this class we have [0.6] 
cytosine [1.0] and uracil [1.3] and as well as adenine in this class we have 
guanine [4.1] and the differences as i said are in the number of substituents 
in the ring [0.5] if you want to know more [0.2] about that [0.3] chapter 
sixteen of Campbell [0.4] is where you look [1.9] so those are the bases [1.4] 
those bases are linked [0.2] to sugars [2.0] deoxyribose [0.2] in the case of D-
N-A [0.3] ribose [0.4] in the case of R-N-A [0.5] both of them sugars 
containing [0.3] five carbons [0.6] one two three four five [0.3] one two three 
four five [0.3] arranged in a five-membered ring [0.5] with one of their 
oxygens [1.1] and then this is linked [0.9] to a phosphate group [0.6] and that 
phosphate group is linked through this carbon here [0.5] the so-called five-
prime carbon [3.3] and as you see [0.5] these nucleotides are linked together 
in a strand [0.2] up to a 
polynucleotide [0.5] by bonds [0.4] between [0.7] the phosphate which is on the 
five-prime [0.5] on the five-prime carbon of this [1.1] sugar [0.6] is linked 
[0.4] to the three-prime carbon [0.5] of the next sugar [0.5] so what we have 
is a chain going sugar phosphate sugar phosphate sugar phosphate sugar 
phosphate sugar phosphate [0.4] and off it we have the bases [0.7] so it's like 
a ladder structure [0.6] sugars and phosphates [0.5] but with bases intervening 
[1.4] so if we [0.4] were to put that back on my simple minded diagram [1.1] 
okay here's base sugar phosphate [0.6] what i would have [0.7] then [0.2] is 
this sugar here [0.6] joining on to another phosphate here [0.8] and then [0.3] 
another sugar [1.1] here [0.4] and another base [0.6] here [1.4] and then this 
sugar [0.3] again joining on [0.9] to another phosphate [1.0] and so on and so 
on [0.9] phosphate sugar phosphate sugar phosphate sugar [4.9] so that in very 
simple terms [1.1] is how you make a nucleotide [0.5] something about the 
nomenclature [2.2] well we found that there are five bases adenine guanine 
thymine cytosine and [0.6] er [0.4] uracil [1.2] in fact [0.2] as nucleotides 
they all change their names just to be very 
awkward [1.6] and [0.2] adenine changes to adenosine [1.2] cytosine to cytidine 
[0.5] guanine to guanosine [0.8] and uracil and thymine [1.0] well there is no 
no uracil changes to uridine and thymine to thymidine [0.3] again this is 
something you don't have to take in now [0.6] perha-, er any book will tell you 
those names [3.9] and we've looked at how to build a polynucleotide we build a 
polynucleotide by joining these units through [0.2] base er through sugar 
phosphate [0.2] linkages [4.4] we've also in talking about nucleotides [0.4] 
discovered some of the differences [0.3] between D-N-A and R-N-A [1.0] we've 
discovered that D-N-A [0.3] has thymine as one of its bases whereas r-, R-N-A 
has uracil [0.3] that D-N-A has deoxyribose [0.5] ra-, where [0.6] er uracil [0.
3] ha-, er where [0.3] R-N-A has ribose [0.7] but there are one or two other [0.
5] structural differences that perhaps we might like to look at [0.9] and one 
of those [6.8] while we're looking at it [5.1] is that D-N-A [0.5] in general 
[1.1] forms a double helix [0.4] of two chains of polynucleotide [1.3] and is 
in general [0.2] a very long polymer [0.5] which has a fibrous structure [1.4] 
whereas R-N-A [0.7] is normally [0.2] consist of one polynucleotide chain it is 
single stranded [0.8] whereas D-N-A is double stranded [0.5] and in general R-N-
A [0.3] chains tend to be [0.2] short [0.6] and [0.2] therefore fold upon 
themselves [0.3] to make globular molecules a bit like proteins [0.9] okay so D-
N-A is a double helix long and fibrous [0.3] R-N-A a single stranded molecule 
[0.3] short [0.2] and globular [2.3] don't know whether you've ever thought [0.
4] how big [0.4] D-N-A might be [0.3] it's a long molecule [1.3] er [1.1] my my 
[0.4] favourite model for it is a [0.6] a piece of [0.8] ordinary sewing cotton 
[0.4] which is a roughly one in a hundred-thousand scale model [0.8] okay it's 
one in it's a hundred-thousand times larger than a piece of D-N-A [1.1] er [0.
7] and on this model [1.1] er the amount of D-N-A in a bacterium [1.0] would be 
a couple of hundred metres [0.2] of this [0.5] okay [0.7] couple of hundred 
metres is actually [0.3] more or less the contents of this er [0.8] this rather 
large [0.8] bit of cotton [0.6] if you can imagine me unravelling all this of 
course you can see what a problem D-N-A is to keep anywhere [0.5] okay [0.7] 
such a 
long molecule clearly although er one draws it conventionally as a nice little 
tiny circle [0.5] is er [0.2] somewhat complex and rather inclined to get 
muddled [0.2] and twisted [0.5] and that's a problem [2.0] for a bacterium as i 
said about two-hundred metres of this stuff [0.4] er [0.7] in each cell in your 
body [1.1] right using this model [0.6] you have a hundred-and-six [0.2] 
kilometres [1.5] okay [0.7] of this stuff [0.3] okay [1.0] it's it's actually 
if the real stuff [0.3] okay so we scale this down a hundred-thousand times [0.
6] is around one-point-six metres [1.7] okay [1.2] one-point-six metres [0.5] 
so it's roughly the height of a less than average man that's me [0.4] er [0.2] 
okay er roughly my height in D-N-A [0.7] if you can imagine [0.2] an a D-N-A 
molecule my height and [0.2] ten-thousand [0.3] hundred-thousand times thinner 
than that [0.7] er that's how much D-N-A you've got inside every one of your 
cells i said a hundred-and-six kilometres didn't i [0.4] it's a hundred-and-six 
miles [1.2] to well it makes it more familiar [0.6] a hundred-and-six so a 
hundred-and-sixty kilometres [0.8] er [0.4] roughly from here to Bristol [0.5] 
so i 
would have to stretch this stuff from here to Bristol [0.3] to model [0.3] the 
D-N-A inside each one of your cells [0.9] so it's very long [0.7] and even 
given [0.5] that inside every one of your cells there are actually forty-some 
chromosomes so th-, it's not [0.7] one molecule [0.6] a hundred-and-six miles 
long [0.6] it's er forty molecules adding up to a hundred-and-six miles [0.9] 
er forty-eight molecules of course [1.0] er [0.8] but er that does give you 
some idea of scale [1.9] that contains [0.4] three- [0.3] thousand- [0.2] 
million [0.9] nucleotides [0.7] okay roughly we believe that [0.3] each [0.3] 
each D-N-A D-N-A [0.6] in one [0.8] haploid amount of D-N-A [0.2] in one of 
your cells is three- [0.3] thousand- [0.2] million nucleotides [1.6] whereas 
for a our poor old friend [0.4] er the er [0.3] E-coli bacterium it's only some 
four-million [0.5] a trifling amount [3.5] just to go back to the molecules for 
a minute [1.4] looking at D-N-A [0.5] you know that D-N-A [0.2] is a double 
helix [1.4] okay there are some pictures of the double helix there [5.0] two 
things about the double helix i think we need to er [0.5] well at least two 
things we need to 
talk about [0.4] one of them is how the double helix is bonded together [1.0] 
there are two sorts of bonds which hold the double helix together [0.8] one of 
those are so-called [0.6] er base pairs [0.8] which are relatively weak bonds 
what we called hydrogen bonds [0.4] which bond across the centre of the 
molecule [1.0] always between adenine and thymine [0.6] and between guanine and 
cytodi-, cytosine [1.3] with two [0.4] bonds between adenine and thymine [0.4] 
and three bonds [0.3] between guanine and cytosine [0.5] that means [0.4] that 
a G-C base pair is roughly fifty per cent stronger [0.4] than a A-T base pair 
[1.7] however that's not the only thing that holds the D-N-A molecule together 
[0.8] as you see from this [0.4] picture [0.4] those D-N-A bases are s-, on top 
of each other [0.3] stacked on top of each other in fact this bit of the 
picture shows you that better [0.6] in this space filling model here we see the 
bases [0.4] stacked on top of each other [1.0] okay those bases being stacked 
on top of each other [0.3] they actually interact with each other [0.5] like 
this [0.3] and that gives an 
extra [0.3] layer of stability to the molecule that's called base stacking [0.
6] so base pairing [0.3] added to base stacking [0.4] stablilizes the double 
helix [2.9] looking at this diagrammatic [0.2] model of the double helix here 
[2.3] some of the details we can count that there are one two three four five 
six seven eight nine ten bases per turn [1.2] off the double helix [0.6] and 
that amounts to a distance [0.4] of some three-point-four nanometres [2.0] and 
the width of the double helix [0.4] is about two nanometres [2.3] a nanometre 
remember [0.3] is one [0.5] times ten-to-the-minus-nine [0.3] of a metre [2.6] 
and the other thing i need to tell you about [0.6] is if we go back [1.3] one 
thing i need to stress and it's one of the mysteries of nucleic acid structure 
[0.6] that always mystifies people [4.3] 
if we [0.4] look at that structure [0.2] as we've draw it here in very simple 
terms [1.1] with our [0.3] sugar [0.3] phosphate sugar phosphate sugar 
phosphate backbone [1.2] okay [0.3] we have at one end of the molecule here [0.
7] a [1.2] something with no phosphate on it [0.8] we conventionally call that 
the three-prime end [0.6] because there's a free three-prime carbon [0.5] stuck 
here [0.7] and at the top here we have a free phosphate [0.8] and that's 
attached to a five-prime carbon here [0.5] so we call that end of the molecule 
the five-prime end [1.1] okay [0.7] so just as proteins have two different ends 
we call them the amino terminus and the carboxy terminus of a protein [0.7] we 
talk about the five-prime and the three-prime end of a nucleic acid [1.4] okay 
[0.9] and if we draw a double helix [1.2] whereas one of the strands is moving 
from five-prime to three-prime in this direction [0.6] the other strand [0.2] 
is moving from five-prime [0.6] to three-prime [0.5] in the opposite direction 
[3.4] that is [1.3] a double helix with its [0.7] bonds in between [1.1] we 
call it an anti [0.6] parallel [0.8] double helix [2.2] 
okay because we have two parallel chains [0.3] one running in one direction [0.
3] and the other one [0.2] running in the other [1.6] and half of the problems 
that people get into thinking about D-N-A [0.3] has got [0.6] to do with 
forgetting [0.3] that they two run in different directions [0.6] that was the 
thing that most people got [0.4] wrong [0.2] for instance in the diagnostic 
test where you were able [0.5] to [0.3] to write me [0.3] the complement of a D-
N-A molecule [0.3] you normally wrote it the wrong way round [0.8] because the 
convention is [0.3] that we always write D-N-A sequences starting at the five-
prime and ending at the three-prime [0.6] so the five-prime is always on the 
left [1.0] my right in this case okay and we put out the sequence [0.3] running 
[0.3] from left to right with the five-prime there [0.4] and the three-prime 
there [4.7] now that's [0.4] a molecule of D-N-A [1.3] what about a molecule of 
R-N-A [0.3] what i just wanted to show you [0.5] as i said they're short [0.4] 
and er [0.5] generally [1.4] globular [1.3] and this picture which you haven't 
got a copy of [0.3] but you can look up in figure [0.4] if you see it's in 
chapter seventeen of Campbell [0.6] it's a picture of a T-R-N-A which we'll be 
talking about [0.3] again [0.5] er in the next er [0.2] part of [0.3] in the 
next talk [1.0] but a T-R-N-A [0.5] is a short molecule [0.6] if you could 
count it's about seventy-five nucleotides long it's trivial compared with the 
[0.5] length that we've been talking about in terms of D-N-A [1.5] okay [0.4] 
and as you see it [0.2] although it's single stranded [0.5] there's only a 
single molecule there it coils back on itself [0.4] making double helical [0.3] 
er making base pairs across here [0.5] and making little bits of double helix 
[0.7] as you can see here [0.5] in this molecule [0.3] making it in fact [0.9] 
almost [0.3] a a little blob [0.2] shape like a protein [0.5] okay [0.6] so 
rather than those enormous great molecules three-thousand-million nucleotides 
long for a piece of D-N-A [0.6] er [0.4] we've got something seventy-five base 
pairs long [0.3] well [0.4] R-N-A molecules get larger than that but not a lot 
larger [2.1] perhaps a few thousand at most [2.2] okay so that's looking at the 
differences between D-N-A and R-N-A [0.9] and while we were looking at 
those differences [1.2] we [0.2] er [0.2] talked a little about [0.8] these 
forces that bound the D-N-A [0.3] together [1.0] okay and you've got that on 
one of your [0.3] overheads [0.5] base pairing [0.4] with hydrogen bonds two 
for an A-T base pair three for a G-C base pair [0.5] and the base stacking [0.
4] bonds which hold [0.3] the structure together [3.5] at this point [0.9] 
we've talked [0.4] almost as much as anyone can bear i think about structure [0.
5] so we'll take a two minute break there [0.8] okay and then we'll go on to 
talk a bit more about the function of D-N-A 
nm0238: 
so D-N-A has to function as the genetic material [0.9] in most [0.4] cells [0.
2] and most organisms [0.8] there are a few organisms very simple organisms who 
use R-N-A as genetic material [0.8] most obviously some viruses [0.6] and some 
rather odd things that plants have called [2.1] but for most organisms D-N-A is 
the genetic material [2.5] and that means that he has to do two things [3.1] it 
has to act as a store [1.5] we have to be able to store information [1.6] and 
we have to be tra-, be able to transmit information [1.6] and there are 
basically two modes of transmitting information [0.6] there's transmitting 
information from one cell to another [1.9] and there's what we call expressing 
information [0.6] that is [0.5] moving from genetic information [0.6] stored [0.
4] to [0.2] phenotype information [0.2] expressed [2.3] which as you remember 
in the case of [0.4] molecules is talking about [0.3] moving [0.3] from D-N-A 
[0.3] to proteins [2.9] so [0.2] D-N-A how does it function as an information 
store we have an 
incredibly long molecule [1.1] a copolymer [1.1] right [0.5] which links 
together [1.5] four different [0.6] nucleotides [1.0] adenine the bases you 
paired your bases adenine guanine cytosine and [0.6] thymine [1.2] A [0.5] G C 
and T as we call them [1.7] okay [1.6] what we have and people [1.1] er [0.8] 
very soon realized this [0.4] is basically [0.4] a [0.3] piece of linear [0.6] 
coded [0.2] information [0.6] and the best analogy that i can think of is 
magnetic tape [1.2] okay [0.4] now you all vaguely know that what we have in 
magnetic tape is a series of iron atoms [0.7] okay which are laid out on this 
plastic tape [0.6] in some sort of [0.2] order [0.8] okay and the m-, [0.2] the 
magnetics the magnetization of those iron atoms [0.7] is [0.3] some indication 
[0.6] of [0.6] information it can be read [0.2] somehow [0.3] by a machine [0.
3] and interpreted as sound [1.0] okay we have a linear [1.9] er [0.4] a linear 
[1.0] succession of these things which we can pull [0.2] so we can [0.3] have 
these sounds in a linear [0.6] er arrangement [0.2] as we like music to be [1.
3] okay [0.2] well you can imagine D-N-A like this [0.3] so that the linear 
sequence of the bases along the strand [1.0] okay [0.6] is is the information 
[0.7] that is the information 
the sequence of bases on a D-N-A strand [1.8] okay [0.7] and exactly like this 
cassette tape [2.0] you neither you nor i [0.8] can tell [0.9] whether [0.5] 
this is Cerys Matthews [0.2] or Dame tiri game Dame [0.2] Kiri Te Kanawa [0.6] 
whether it's [0.2] Beethoven [0.6] or [0.3] er Catatonia [1.7] er [1.3] because 
[0.3] just like this [0.4] the information is [0.2] redundant [0.2] useless [0.
4] okay [0.8] this information is no good unless interpreted [0.3] by some sort 
of interpreter which in the case of a cassette tape of course [0.5] is a 
cassette player [0.9] but in the case [0.3] of [0.2] D-N-A genetic information 
is the cell [1.4] okay [0.4] so that information [0.2] only has sense in the 
context of the cell in which it finds itself [1.4] okay and if you want to 
think mechanistically [0.2] you could think of a cell [0.3] as a cassette 
player [0.4] but it's not really helpful [0.2] i don't think mechanistic [0.3] 
analogies really work [0.7] okay [0.2] but something needs to interpret that 
genetic information that clearly is the cell [1.8] so the information store 
consists of the sequence [0.5] of the bases [0.2] in the chain [1.5] the 
transmission of information as i see it happens in two 
ways [1.3] first of all i'm going to talk about [0.2] transmission by [0.2] 
expression [0.6] and then we'll talk [0.2] later [0.5] about what i've called 
on this slide [0.4] maintenance [0.3] of [0.2] information but it's also 
transmission to the next generation [0.9] okay [0.3] and its replication [3.0] 
in order to [0.2] look at [0.2] transmission of information i'm going to go [0.
4] to what [0.7] i've shown you i think once already [0.4] which is what [0.2] 
er [0.9] Francis Crick immodestly called the central dogma of molecular biology 
[0.8] this little diagram [0.5] which shows you the information flow inside a 
cell [0.5] from the D-N-A store [0.6] of genetic information [0.7] through a 
temporary [0.9] resting place in R-N-A [0.4] particularly messenger R-N-A [0.9] 
to the protein [0.3] which is equivalent to the phenotype so from genotype [0.
4] to phenotype within a cell [1.0] when [0.5] Crick [0.2] wrote this out first 
of all [0.3] he wrote that diagram with simple arrows on it [0.5] showing that 
the information moved from D-N-A to R-N-A to protein [2.1] okay [0.2] and 
that's equivalent to the move from genotype to phenotype [2.6] now that's [0.3] 
although [0.3] Crick would [0.3] deny it heartily 
that's actually a very simplified form of the central dogma [0.4] and we'll 
look [0.4] at a more complex form in the next lecture [1.1] okay [0.3] but 
that's basically [0.2] how the information was transmitted [0.5] inside the 
cell [0.5] to interpret [0.3] the ph-, genotype as a phenotype [2.0] what i 
want to [0.6] concentrate on [0.5] in this lecture [0.6] is the next stage [0.
5] which is replication [0.8] what the cell has to do [3.2] because that cell 
is going to divide [0.2] and have daughter cells [0.3] what that cell needs to 
do [0.2] in order to pass on that information to maintain [0.6] a decent store 
of information [0.6] in its daughter cells [2.8] one of the big differences 
between these two processes transmission [0.7] and maintenance [0.8] is that it 
is much more important [0.4] for this one to be accurate [0.4] than it is for 
this one to be accurate [1.3] okay [0.5] when we're trying to maintain the 
information [0.3] every time we copy the information if there are mistakes [0.
8] introduced [0.4] then we've got mutations [0.4] and we've got a change in 
our genetic information [0.8] okay so we want to make absolutely sure if we can 
that replication [0.2] is as accurate as be made [0.9] with transmission of 
information moving information within a cell [0.4] from [0.2] our D-N-A to R-N-
A to protein [0.3] we're not so worried [0.2] because we're going to throw that 
R-N-A and that protein away at the end of the day [0.2] we're going to make new 
copies [0.3] and those new copies may be more accurate [1.8] so although 
accuracy is the important consideration [0.3] for both transmission and 
maintenance it's really the prime [0.4] problem [0.3] for maintenance [2.4] so 
the process [0.5] whereby we take [0.2] a D-N-A double helix and make two D-N-A 
double helices [0.4] is the process we refer to as replication [0.5] and that's 
what i want to spend the rest of this lecture talking about [4.6] 
and D-N-A replication [1.3] in very simple terms [0.2] has four important 
characteristics [1.5] the first is that it is semiconservative [1.7] what's 
that mean [0.7] one usually makes a political joke here but i won't make a 
political joke [0.5] er you can work out a political joke for yourself [1.7] 
semiconservative means [0.3] that if you take a D-N-A double helix [0.3] and 
here i'm going to [0.4] turn to my other favourite model [0.3] of a D-N-A 
double helix which is a piece of Velcro [2.0] okay [0.5] the important thing 
about a piece of Velcro is as you know is that it has two sides [0.3] which 
better bind to each [0.3] themselves [0.2] but which bind to each other [0.5] 
this entirely mimics [0.3] the fact that [0.2] one [0.8] chain of a D-N-A 
double helix [0.3] absolutely specifies and will only bind to er the other 
chain [1.4] okay [0.4] and we say that the chains of a D-N-A double helix are 
complementary [0.3] to each other [0.4] 
that's complementary with a E [1.1] C-O-M-P-L- [0.3] E- [0.4] M-E-N-T-R-Y-T-A-R-
Y [0.5] okay they're complementary to each other [1.2] right [0.6] we can 
describe that in very crude terms and say they stick to each other [0.4] and 
some people [0.3] and we do in molecular biology refer to these strands as 
sticky [1.2] okay [1.4] but we've got [0.5] here's our double helix it's not 
helical but you could imagine it [0.7] what happens [1.0] do we part this 
double helix [1.1] make a new chain on here and make a new chain on here and 
join it up again [0.9] which would be a conservative model [0.2] because this 
helix would remain exactly the same [1.7] okay [1.0] or do we [0.2] throw away 
both strands of this helix and make a new one well that would be wasteful and 
clearly we don't do that [0.6] what we do do [0.2] is semiconservative 
replication that is we take these two strands [0.5] and on each of these two 
strands we build a new one [0.9] so for each strand [0.5] half of it is old 
there's half of it new [0.2] and hence semiconservative [2.9] and that's very 
clearly shown in the diagram which i think you've got a 
version of [0.5] on your handout [1.3] okay [3.6] this as i said will be a 
conservative model where we started with a double helix [0.6] when we made a 
new double helix that was entirely new and the old one was kept [1.6] right [1.
0] that's a conservative model [0.5] the semiconservative model is where we 
start with a double helix [0.3] we part the two strands and make a new one from 
each [0.6] and again [0.6] when we make that one we part the two strands and [0.
3] and er make a new one on each [0.2] don't worry about the dispersive model i 
don't think it's worth [0.7] thinking about at the moment [1.5] okay [0.7] so 
we say that D-N-A [0.3] synthesis is semiconservative what evidence do we have 
for that [0.9] the evidence that we have for that [0.4] is the evidence of the 
Meselson-Stahl experiment [0.6] which again you have [0.3] on your handout [6.
6] okay this was an experiment which was done [0.8] in er the nineteen-fifties 
[1.0] er [0.7] in which what the er [0.3] the two [0.4] authors Meselson and 
Stahl did [0.8] was to label [0.6] the strands of D-N-A this time in the 
Hershey-Chase experiment remember we s-, [0.3] labelled D-N-A [0.4] and 
labelled protein [0.8] in the men-, [0.2] Meselson-Stahl experiment what we do 
[0.3] what we did [0.6] was to label the old strands of D-N-A [0.3] with one 
isotype [0.2] not a radioactive one in this case [0.7] of nitrogen [0.8] okay 
and the new strands with the new isotope [0.8] okay [1.4] and [1.1] we then [0.
2] look at [0.7] the [0.3] D-N-A molecules that were formed [0.6] in fact what 
what they did [0.6] was to start [0.5] with nitrogen fifteen [0.9] and then 
they substituted it with nitrogen fourteen [0.5] nitrogen fourteen is lighter 
than nitrogen fifteen [0.3] so the density of the molecule went [0.4] decreased 
[1.3] and they showed that density [0.4] by [0.2] running [0.6] the D-N-A 
molecules in an ultracentrifuge [0.9] so here's what they started with in the 
ultracentrifuge this is heavy D-N-A [1.3] okay [0.8] and what you see happening 
[1.0] is that as we run [0.3] through one [0.4] cell cycle [1.1] the D-N-A 
moves from this position [0.8] to all be at a new position here [1.2] okay [0.
3] and as we run through a second cell cycle [0.3] it moves to being at two 
positions [2.1] if we go back to our model [2.3] we'll see that that's exactly 
what we 
expect [0.9] from our semiconservative model [1.1] we start [0.5] with dark 
blue heavy [0.8] and we move through in one [0.3] cell cycle [0.3] to 
everything being a mixture of light an-, and dark blue [0.3] that is the same 
density [0.5] whereas in the conservative model [0.3] we would move [0.3] from 
blue [0.7] to [0.4] dark blue and light blue two different [0.4] densities [0.
8] so at the end of the first generation you have two different densities [0.5] 
whereas at the end of the first generation here we [0.3] just have one density 
[0.8] at the end of the second generation [0.5] however in both cases [0.6] we 
will have two densities [1.2] okay [1.2] ar-, in conservative [0.5] three of 
them will be light one of them heavy [0.5] in the case [0.2] of 
semiconservative two of them [0.5] will be mixtures [0.3] and one of [0.2] er 
two of them light [0.7] and if you again look at the Meselson-Stahl experiment 
[0.6] you'll see that that's what happens in the second generation [0.4] we 
move from this position here [0.3] to a mixture of things of that density and 
some lighter things [0.6] okay [0.6] so by measuring the density [0.6] of D-N-A 
[0.3] during the experiment in which we 
substituted [0.4] a light nitrogen isotope for a heavy nitrogen isotope [0.5] 
we were able to prove or Meselson-Stahl were able to prove [0.3] that 
semiconservative [0.2] was the correct model [0.4] for D-N-A replication [2.8] 
what other characteristics [0.5] do we need to know about D-N-A replication 
well first of all [0.8] does it go all over the place no it doesn't it always 
starts in one place [0.6] starts at what we call an origin [1.5] and in the 
case of er a bacterial [0.4] chromosome there is only one origin [0.9] okay 
there's only one place where we start and replicate bacterial chromosome [0.7] 
in the case of your and our my chromosomes which are infant as we saw [0.2] a 
thousand times or so larger [1.2] er [0.4] i'm afraid we don't start from one 
origin but we have many origins otherwise it would take us forever to replicate 
our D-N-A [2.9] however from that origin [0.5] we can very easily show [0.3] 
that the D-N-A r-, is replicated in both directions [1.0] okay [0.2] that 
replication is bidirectional [1.8] so if we imagine [0.4] that we're going to 
start making a piece of D-N-A [2.5] 
so here i have a piece of D-N-A here's a double [0.7] strand of D-N-A [0.7] and 
here is the origin [0.8] okay [0.6] obviously the first thing that must happen 
at the origin is w-, me must [0.5] dissociate the strands like that [1.3] and 
you can in fact see that happening in an E-M photograph a little bubble will 
appear [1.5] in the D-N-A [0.8] okay and then we start making D-N-A in both 
directions that is we make it on this strand [0.8] okay and we make it on that 
strand [2.8] and D-N-A is always made in the same direction [1.0] it's always 
made [0.2] starting [0.2] with the five-prime of the new strand and moving 
towards the three-prime of the new strand [1.3] which since those two strands 
[0.2] as i said are antiparallel [1.2] this strand has a three-prime here and a 
five-prime there this strand has a three-prime there and a five-prime there [1.
9] okay [0.4] those two directions will be the opposite directions [1.7] here 
we have [0.9] the snag [0.7] of D-N-A replication [0.7] because unless [0.6] 
when i replicate bidirectionally i'm going to be satisfied [0.7] with going 
this way all 
the way round my circular D-N-A [0.2] if i'm a bacterium [0.8] okay and come 
back to meet myself here which will take me a long time [0.5] and do the same 
in the opposite direction for the other strand [1.2] i'm [0.2] i'm [0.6] if i'm 
satisifed with that it will be a very long process [0.6] and also [0.2] i will 
have a single stranded piece of D-N-A hanging around for an awfully long time 
[0.5] which you can imagine what it looks like like this you can imagine the [0.
2] problems you'd get into [0.9] er [1.1] we need a way of replicating the 
other strand at the same time [2.0] and as i've told you [0.9] we can only 
replicate D-N-A in one direction [1.1] and the solution that D-N-A has [1.7] is 
that although [0.3] it replicates D-N-A continuously on one strand [0.5] it 
reputat-, [0.2] replicates the other strand [0.3] discontinuously [1.9] okay [1.
6] and that means we make the other strand in fragments [2.7] so if you imagine 
the other strand being made [0.5] what i do [0.4] is to start here [0.7] and 
make a little bit of D-N-A [0.8] and then i jump back and i start here and make 
another little bit of D-N-A [0.5] and then i jump back and 
start here and make another bit of D-N-A and so on [1.6] a motion i've 
described and it will only mean something to the [0.3] girls among you i guess 
as being something like blanket stitch [1.1] okay in out round the back in out 
round the back [0.4] okay [1.1] those of you who think i've got the 
nomenclature of the stitch wrong can come up and tell me later but [0.5] i 
assure you there is a stitch [1.7] okay [1.1] but you see we're going forwards 
[0.3] the general direction of synthesis is going in this direction [1.1] okay 
but i'm always actually making the molecule in this direction [0.5] that is the 
five-prime to three-prime direction [0.7] i clearly i'm going to make a lot of 
fragments [1.3] and those fragments have a special name they're named [0.5] 
after the Japanese scientist [0.6] who first discovered them [0.5] they're 
named [0.3] Okazaki fragments [4.1] so D-N-A replication is [1.1] 
semiconservative [0.7] we retain one old strand in each new double helix [0.7] 
it starts from a single place of origin of replication [0.9] it moves in both 
directions from that origin [0.8] moving continuously on one 
strand [0.3] and discontinuously [0.4] on the other [3.2] of course what's 
happening [1.2] on that strand [0.2] is something very simple [1.4] okay and 
the basic mechanism of D-N-A replication is very simple [0.8] we have [0.2] a 
double stranded helix [0.4] we part it [0.7] and then we simply match [0.6] new 
bases [0.8] to replace the new strand so where there's an A we put a T where 
there's a C we put a G [0.4] and so on [0.5] until we get two new strands [0.4] 
we're matching [0.4] bases [4.6] how [0.2] do we do that [0.6] how do we do the 
whole thing what is the mechanism of D-N-A replication [4.0] and that's where 
[0.8] where i want to stop with thinking us through the mechanism of D-N-A 
replication [0.8] and we'll then look at the diagram that you've got [0.4] 
lastly on your handout which illustrates it [0.9] so firstly [0.2] obviously [0.
2] we need to unwind that double helix we need to make our replication bubble 
as we sometimes call it [4.0] nature uses two mechanisms to underwi-, unwinding 
the double helix it uses an enzyme which actually physically unwinds it [0.6] 
it's called a helicase [1.3] okay and it actually does that by untwisting [0.4] 
the Watson-Crick double helix [2.0] and it uses the device of having a protein 
in the cells [0.4] which binds strongly to single stranded D-N-A but not to [0.
3] double stranded D-N-A at all a single stranded binding protein [0.7] okay so 
as soon as we part the strands [0.5] this protein binds hard [0.3] and the [0.
2] the strands are disinclined to come back together [0.5] then we use two 
mechanisms [0.3] for keeping the strands apart [3.4] then we need to start 
making some D-N-A [0.9] and we have a problem [1.1] we have a problem [1.3] 
that problem is [2.1] clearly we need [0.2] to get [0.3] our [3.5] mechanism 
right so that we get the right base pairs [1.2] and the best way to get the 
right base pairs matching [1.0] is to have an enzyme [0.9] which knows [0.2] 
roughly [0.7] how wide a D-N-A double helix is going to be [0.5] and can fit it 
[0.4] okay fit [0.2] to the helix we've got [0.2] fit to the distance [0.4] 
because an A-T base pair and a C-G base pair are the same size as each other [1.
2] okay [1.6] and [0.6] the enzyme can't do that [0.3] at the same time [0.5] 
as starting a strand on its own [1.8] the enzyme which makes D-N-A [0.2] will 
not start a strand on its own [0.9] so it needs to be helped [0.9] and it needs 
to build a primer first [1.5] okay [0.6] and that primer's always made of R-N-A 
[1.3] 
okay so we have to first in order to make D-N-A make some R-N-A [2.1] and what 
this means basically is going to this [0.7] going over here [0.6] let's imagine 
we have our [0.6] piece of D-N-A [1.0] okay [0.3] imagine this is the five-
prime of it and this is the three-prime of it and we're going to make [1.0] er 
[1.0] some D-N-A here we're at the origin [0.7] we start with a piece of R-N-A 
[0.9] which [1.0] bonds like that [0.9] okay made from five-prime to three-
prime [1.7] and then the enzyme which makes D-N-A takes over from this point [0.
7] and makes D-N-A [0.4] in the right direction [1.5] okay [1.7] so at the 
beginning [1.2] of [0.2] our [0.2] D-N-A we have [0.3] a combination of a 
primer made of R-N-A [1.7] and an old strand [0.4] of D-N-A which we're using 
[0.2] to read [0.4] the correct sequence which we call a template [0.9] okay so 
at the beginning we have a template and a primer [6.4] so we start the chain 
with R-N-A [0.2] we make a primer [0.4] and then we continue D-N-A synthesis [0.
9] on the continuous strand that we call 
the leading strand [1.2] okay because it's always slightly ahead [0.2] of the 
other strand in making D-N-A [1.3] okay [0.3] and on the other strand the 
lagging strand as we call it 'cause it is slightly behind [0.7] okay [0.7] we 
make those fragments those Okazaki fragments and we need one more process to go 
on and that is we need to join the fragments together [1.2] okay so we need a 
ligase [0.3] a joining together enzyme [0.3] to join the fragments together [2.
0] so we can incorporate all that information about the mechanism of 
replication [0.6] on the diagram here [0.6] that i've given you a copy of [2.4] 
okay which gives you a brief summary [0.4] of D-N-A replication here's the 
parental D-N-A which has started [0.7] it's being unwound at this point [0.7] 
and there are single stranded binding proteins these are the pink fingers [0.6] 
binding to the single stranded D-N-A [2.1] all right [0.2] then we [1.0] we 
have D-N-A being made [0.4] continuously in this direction [0.9] okay by the 
enzyme 
which makes D-N-A the D-N-A polymerase [1.3] and on this [0.3] strand [0.4] 
we're making bits of D-N-A [1.7] but starting with R-N-A primers [0.6] to make 
our Okazaki fragments [0.6] we're then extending our Okazaki fragments [0.9] 
and in the end [0.6] here's the D-N-A ligase enzyme joining two Okazaki 
fragments together [1.1] so we unwind [0.9] continuously make D-N-A on this 
strand [0.5] five-prime to three-prime direction [0.4] we discontinuously make 
it on this strand starting with an R-N-A primer [1.5] and then making our D-N-A 
until it bumps into the next one basically it just falls off and it bumps in [0.
8] and then we have to join up our two Okazaki fragments [0.3] using a ligase 
[1.1] so that is the basic mechanism [0.3] of D-N-A replication and that's 
where we shall end [0.6] this first [0.5] er [1.1] talk for this afternoon [0.
7] okay [0.3] i'll be putting out some more handouts for you for the second 
talk in just a second [0.8] we'll take a five minute break