Slide. Short segment of a DNA molecule. 20 bp This is, of course, the fundamental structure from which and around which contemporary
biologists think about living things.
The totality of the DNA in a cell, be it a single independent cell that is itself and organism (bacteria, yeast) or one of
billions of cells in a complex multi-cellular organism (man, corn plant) is called the genome.
Mr. Pres. Fellow members: - gratitude
The students of the late 1980's take this DNA molecule and the major biological principles that flow from it for granted.
But it was only in the late 1940's and early 1950's, when I was an undergraduate and still thought that I wanted to
be a chemist, that the first of these principles was established.
Avery, then Hershey and Chase, established that genes, genomes, the informational systems that encode the competencies of
living things resides in DNA.
I was only vaguely aware of this notion as I took the first step toward biology -- entering graduate studies in biochemistry.
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While I was a graduate student, busy with enzymology and phosphoproteins, the second gigantic step was taken. Watson and
Crick proposed the double-helical, vase paired structure for DNA. There was no way to avoid being challenged by the intellectual
and experimental attractions of this macromolecule and its close relative, RNA.
Upon completion of the Ph.D. degree I entered the world of nucleic acids by becoming a post-doctorate fellow with Leon Heppel
at the NIH. The year was 1956.
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It was a small world, a few laboratories scattered about the globe. For me , it was the start of 30 years of unflagging scientific
excitement and the start of association with a group of people that have become my most cherished colleagues, and friends.
These days when, in a years time, 1000's of people attend 10's of meetings on nucleic acids each year, it is amusing
to remember that in the late 50's, the nucleic acids group was given one morning, grudgingly by those who ran the week
long Gordon Conference on proteins.
The central contribution of the mid-to late-fifties -- was the recognition that enzymes are the key to dealing with nucleic
acid structure, although we were all unsuspecting of just how versatile in reaction and specificity such enzymes would prove
My own work in those post-doctoral years an in the first decade of independent scientific work was to study the mechanism
of the synthesis of polyribonucleotides (RNA if you will) by PN Pase, the enzyme discovered first polymers[?] in the mid-fifties
by S. Ochoa and M. Grunberg-Manago. By 1960, I had learned how to make a variety of polyribonucleotides. For that reason
when the 3rd major principle of modern biology was established I could be a participant, not a distant admirer.
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In efforts to establish the nature of the genetic code. 3 base pairs specifies a codon on 1 amino acid. Including 1 codon
"start". 3 codons "end"
Along with several other NIH scientists I put aside my own experiments to assist our colleague Marshall Nirenberg. My own
contribution was to prepare RNAs of different structures. We thought it was hi-tech work. From out current vantage point
it seems almost trivial, embarrassing even to mention.
My interest in the mechanism of polymerization and depolymerization continued through the 60's. Along with the first
post-doctoral fellows to join my group, I demonstrated that enzymatic synthesis degrading of long polymers can occur by what
we called a processive mechanism. Processivity is now known to be quite common. Consider the following: and enzyme starts
the synthesis of a new polymer by catalyzing the joining of the first 2 monomer units. There is then a choice. First the
enzyme may drop this down[?] and then [ . . . ] synthesize another, or add a third unit to the original. After each addition,
the enzyme continues to release the product and then pick randomly between extensions of existing chains or starting a new
one. In this model, all the new chains grow together one time. In the second, the processive model, the enzyme hangs on
to the new chain once it is started and adds to it new monomer units. When the chain grows long enough, the enzyme drops
it and imitates synthesis of another chain. Here, there are a few very long molecules even very early in the reaction.
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Completely symmetrical arguments can be made for the degradation of chains by processive or random mechanisms.
The mechanistic work on polynucleotide phosphorylase and bacterial rubonucleases was rewarding, but my frustration grew as
approach after approach failed to define the functional role of these enzymes in bacteria. It was time to shift gears. This
change was initiated with a sabbatical year at the Weizmann Institute -- where I spent the time learning about mammalian cells
and viruses whose genomes are DNA. Returning to the NIH, I spent the next few years trying out various ideas and systems.
During this time the methods we now refer to as 'recombinant DNA techniques' or 'genetic engineering' emerged.
Mention: complete dependencies on enzymes as reagents. And as their extraordinary capacity to bear fruit with previously
intractable problems became increasingly clear, it proved impossible if not foolish to resist their seduction. Especially
because they seemed applicable to a truly peculiar puzzle, a puzzle important enough to be given a name. The c-value paradox.
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The paradox is simple to describe.
Average size of a gene - 1200 bp (400 codons).
SLIDE. E.coli (bacteria), 4000 genes 6*10^6 bp.
Complex multi-cellular. (fly, mammal), [ . . . ]
What is all the extra DNA?
Repeated DNA sequences. Segments 2bp = 10^3 where more or less the same order of bases is repeated. Rec. DNA techniques
have permitted us to begin to analyze(evolutionary implec[?]) First step: classification. High, middle, [ . . . ]
Interested in highly-repeated > 104
SLIDE: Tandem and[?] Interspersed
SLIDE: Types of repeats
SLIDE: Amounts of repeats
Our most recent efforts -- with Lines. Line 1
SLIDE: Summary of Line 1 in all mammals
SLIDE: Structure of the longest lines. Define pseudogenes implies genes
SLIDE: Major Questions.
SLIDE: Mechanism of pseudogen formation. RNA -- DNA various problems: