DNA

module not yet titled

electrophoresis of DNA and proteins

the Sanger method of DNA sequencing

LINKAGE MAPPING

In humans, 1 cM is roughly 1 million nucleotide pairs (bp) of DNA.

The future of genetic research

year the idea of defining an organism by the sequence of its DNA bases

Large Molecules

Diagram Structure of DNA and RNA

DNA Structure

Control of the Cell Cycle

DNA together with associated proteins. This process of doubling the

The period during which DNA replication occurs is not spread

DNA replication is accomplished during a discrete window of time, termed

hours, during which time the entire complement of chromosomal DNA is

completion of DNA synthesis and chromosomal replication in S phase,

Use of DNA in Identification

Use of DNA in Identification

Use of DNA in Identification

Dr. Eric S. Lander at the "Winding Your Way through DNA" symposium,

in DNA coding among human beings. . . .

in this audience who has the same DNA sequence as anyone else.

And indeed, your DNA sequence is unique amongst all DNA sequences of

someone who has the same DNA sequence. But apart from that, your DNA

DNA sequence behind here in some form in some biological tissue, in

Thus is born the notion of DNA identification. And it was quickly

realized that this DNA identification would be especially useful in

until it was possible to read DNA.

DNA gives us rich results, and just as detailed as a fingerprint, in

out a DNA text in its entirety. It would be a wonderful thing if we

we do DNA comparisons, we can't read all three billion letters. What is

By taking that DNA and cutting it with an enzyme that recognizes a

radioactive DNA from this region, one can visualize bands

site, and the next site, and compare it to the DNA patterns taken from

If Suspect #1 has a different DNA pattern than the evidence, the

relates to the crime. But that evidence sample of DNA cannot possibly

Suspect #2's DNA corresponds perfectly at each of the four places of

Those are simply the ideas underlying DNA fingerprinting, as it's

popularly called, or DNA typing or DNA identification,as we prefer to

call it. Within five years of the notion of DNA spelling differences

Selmar, Lifecodes, and others, which grew up to provide DNA typing

DNA typing lab in the Hoover Building in Washington. There were dramatic

ad). "DNA Fingerprinting Links the Criminal to the Crime," with the

typically, about mistaken identity being the problem, because DNA from

immediately upon DNA testing. Many rapists, because of this, now

In essence, DNA evidence is rapidly becoming, in principle, an

done right. Fights erupt over DNA fingerprinting

well-regulated the practice is. For example, DNA fingerprints should

of a DNA fingerprint, which was used in a criminal case in New York.

good. It was the first case in which DNA fingerprinting was actually

of the different DNA patterns of different genes vary across the

Similar things are known for other types of DNA differences. And so

gestation, of an NRC report called, "DNA Technology in Forensic Science."

for the laboratory practice of someone who will create a DNA fingerprint

creating national databases of everyone's DNA type. That way, when a

segment sample and get its DNA pattern, and compare it to a database of

everyone's DNA pattern and find out whose it was. There are many people

is a marvelous technology to amplify DNA. It allows you to take a

specific region of DNA on the chromosome, and by using little black

in principle it is possible to start from the DNA of a single cell and

get enough DNA to analyze it. That makes it possible not just to

could get one microgram, one millionth of a microgram of DNA, or semen

standard techniques, but in fact even shed hair has enough DNA at its

root. A urine sample, saliva sample, will have enough DNA in most

DNA to trace from the seal of the envelope.

sneeze on something, my DNA is there, too. And so there is tremendous

at times kidnapped and blindfolded from the streets of Buenos Aires,

markers, but this was not terribly powerful in these cases. DNA

powerful, unique sequences of DNA would be needed. And so Dr. King's

group turned to looking at a particular bit of DNA called the

mitochondria DNA. It exists in a little organelle - a little package

outside the nucleus of the cell. It's a small bit of DNA, and what's

read snippets of unique, variable sequence mitochondrial DNA, I can

kidnapped by the military. She was tortured and eventually released.

the courts do DNA testing. Mitochondrial DNA sequences were obtained

Let me also mention to you two other applications of DNA identification

but plants as well. One of the great uses of DNA fingerprinting turns

in fact, now you can do DNA fingerprinting on corn plants, and many

large seed companies routinely maintain databases of the DNA

And then, very briefly, the ultimate in DNA paternity testing. As I

York Times a story about DNA taken from a 40-million-year-old termite,

preserved in amber. And DNA sequencing by PCR has been done on this

DNA paternity testing way back, it brings us to our common origins as

consequences. No one sets out to develop DNA identification, the

looking at DNA spelling differences for the purpose of criminology

DNA spelling differences has had consequences in all of these areas.

Excerpted from the "Winding Your Way through DNA" symposium

An Interview with DNA Forensics Authority Dr. Bruce Weir

An Interview with DNA Forensics Authority Dr. Bruce Weir

An Interview with DNA Forensics Authority Dr. Bruce Weir

The term "DNA fingerprinting" was coined by British geneticist Alec

Jeffreys only ten years ago. Since that time, DNA forensics has become

an important tool in law enforcement. In some cases, the DNA tests have

cases have put the spotlight on DNA forensics and created the impression

controversies surrounding DNA evidence.

mean when we say DNA fingerprinting?

A: DNA fingerprinting, or DNA profiling, as I prefer to

call it, characterizes a small portion of our DNA. It is a way of

identifying the DNA content of an individual. We think of fingerprints

identical twins have different fingerprints. In contrast, DNA typing,

because it uses a very small fraction of the DNA, is certainly not

involved. It is also possible to extract DNA from exotic things like

they may obtain DNA samples from a skeleton.

crime scene be commingled with all kinds of other DNA from bacteria,

flora, etc.? How is this extraneous DNA excluded?

A: The methods used in forensic DNA are not so much

concerned with excluding extraneous DNA as with the identification of

human DNA. The probes used are very specific for human DNA.

forensic DNA testing. Are the laboratories using PCR to look at

the RFLP method is that it requires a relatively large amount of DNA

techniques offer the advantage of requiring only trace amounts of DNA,

frequency of the pattern, we rely on a statistical model. A DNA profile

would not overstate the strength of the evidence. The DNA databases

laboratory work would be the weak link in the chain of DNA forensics?

to confirm conclusions, etc. One criticism of forensic DNA profiling as

techniques in DNA profiling. Is one RFLP the same as the next? What

specificity in DNA profiling would be a combination of RFLP plus a PCR?

much debate on the validity of DNA forensics and the statistical

accuracy of DNA profiling?

about and examined the issues carefully who remain critical of DNA

module not yet titled

genomic DNA vs.cDNA

repetitive DNA in eukaryotes

Examples of Viral Replication Pathways

the recombinant DNA is replicated, it passes the viral DNA onto all of

7.01Recombinant DNA Practice Problem

7.01Recombinant DNA Practice Problem

7.01Recombinant DNA Practice Problem

When you do a ligation with a cut vector and insert DNA, there

Solutions to Recombinant DNA Practice Problem

a) So that the DNA can be replicated, so that the daughter cells will

up DNA. Ampicillin kills the un-transformed (and therefore uninteresting) ones.

d) i) If you insert DNA into the EcoRI site, it will add sequence within

Basic Virus Structure

DNA or RNA . A number of viruses contain their genetic information in RNA

instead of DNA). Viruses survive and reproduce by infecting a cell and

The DNA is injected into the bacteria through the baseplate.

Biological Macromolecules

nucleotides found in DNA and RNA

Structure and Function of Organelles

This is where the DNA is kept and RNA is transcribed. RNA is

supporting evidence. Mitochondria have their own DNA and their own

Recombinant DNA Chapter Directory

Recombinant DNA Chapter Directory

Recombinant DNA Chapter Directory

Southerns, Westerns, and Northerns: Molecular Cloning Techniques

Theory: Complementarity

Basic Definitions

Overview

Gel

Transfer to Solid

Blocking

Preparing the

Hybridization

Washing

Detecting of

Summary of Blots

Cloning a Gene By Hybridization

DNA Fingerprinting

DNA Fingerprinting in Human Health and Society

An Interview with DNA Forensics Authority Dr. Bruce Weir

Use of DNA in Identification: A talk by Eric Lander

7.012 Cloning Project: Agricultural Biotechnology

b) Using this data, design a recombinant DNA strategy to render plants

The sequence of the insert DNA has revealed 3 long open reading frames

1) Construct a plasmid library of wild-type bacterium Y DNA in

3) The only colonies to grow would contain a plasmid with insert DNA

Hypertextbook Chapters

Recombinant DNA

Lwoff's Pathways - Viral Replication

called the lytic pathway. They enter and inject a host cell with DNA,

inject their DNA into the host cell - but instead of taking over the

host cell and using it to make viruses, the injected DNA can become

the recombinant DNA is replicated, it passes the viral DNA onto all of

DNA Fingerprinting in Human Health and Society

DNA Fingerprinting in Human Health and Society

DNA Fingerprinting in Human Health and Society

during the 1930s, each person has a unique DNA fingerprint. Unlike a

altered by surgery, a DNA fingerprint is the same for every cell,

treatment. Consequently, DNA fingerprinting is rapidly becoming the

An additional application of DNA fingerprint technology is the diagnosis

The Structure of DNA

also have different DNA sequences. The more varied the organisms, the

more varied the DNA sequences. DNA fingerprinting is a very quick way

to compare the DNA sequences of any two living organisms.

Making DNA Fingerprints

DNA fingerprinting is a laboratory procedure that requires six steps:

1: Isolation of DNA.

DNA must be recovered from the cells or tissues of the body. Only a

example, the amount of DNA found at the root of one hair is usually

Special enzymes called restriction enzymes are used to cut the DNA at

bacteria, will cut DNA only when the sequence GAATTC occurs. The DNA

electrophoresis. The DNA pieces are passed through a gel made from

3: Transfer of DNA to nylon.

The distribution of DNA pieces is transferred to a nylon sheet by

pattern called the DNA fingerprint. Each probe typically sticks in only

6: DNA fingerprint.

The final DNA fingerprint is built by using several probes (5-10 or

Uses of DNA Fingerprints

DNA fingerprints are useful in several applications of human health care

DNA fingerprinting is used to diagnose inherited disorders in both

programs, genetic counselors use DNA fingerprint information to help

other programs, prospective parents use DNA fingerprint information in

depend on the information contained in DNA fingerprints. By studying

the DNA fingerprints of relatives who have a history of some particular

disorder, it is possible to identify DNA patterns associated with the

FBI and police labs around the U.S. have begun to use DNA fingerprints

hundreds of cases have been decided with the assistance of DNA

Another important use of DNA fingerprints in the court system is to

applications, DNA fingerprints bring an unprecedented, nearly perfect

Because every organ or tissue of an individual contains the same DNA

collect DNA fingerprints from all personnel for use later, in case they

are needed to identify casualties or persons missing in action. The DNA

"DNA fingerprints witness for the prosecution." Discover. June 1988, p. 44.

DNA Identity Testing Information Package. Available from LifeCodes,

Genetic Witness -- Forensic Uses of DNA Tests. U.S. Office of

module not yet titled

nucleotides in DNA and RNA

the discovery of the double helical structure of DNA

other key elements of the structure of DNA and its implications for DNA replication and RNA transcription

alternative structures of DNA

the significance of the Messelson and Stahl experiment in the determination of how DNA replicates

the enzymes involved with DNA replication, RNA transcription, and protein translation

Central Dogma Directory

The Identification of DNA as the Biochemical Material

DNA Replication

Cloning Genes

Cloning Genes

another. In this experiment, the DNA of one microorganism recombined

with the inserted DNA sequence of another, and thus had been edited to

The methods used in rDNA technology are fairly simple. We take, for

it into the DNA of Escherichia coli, a bacterium that inhabits the human

of copies of themselves, and each bacterium carries in its DNA a

DNA and paste, or splice, it into plasmid DNA, a special kind of DNA

in a very precise way a specific base sequence of the DNA molecule. With

the human DNA molecule that specifies insulin production can be

This segment is "glued" into place using an enzyme called DNA

ligase. The result is an edited, or recombinant, DNA molecule. When this

recombinant plasmid DNA is inserted into E. coli, the cell will be able

This highly simplified description of rDNA technology does not fully

genetic processes. But we can begin to understand how, by using rDNA, it

7.012 Genetics Supplementary Handout Page 1

DNA.

where l and n are,0.05cM apart. (In Drosophila, 0.05cM is roughly 25000 bp of DNA.

Prokaryotic Gene Regulation

'lac gene' would have been defined as the minimum piece of DNA that

could function on its own - that is, the smallest DNA fragment that

subject to regulation - an mRNA copy is made from the DNA and

these are regions of DNA that code for proteins. These genes produce

DNA sites -

these are regions of DNA which are involved in gene expression or

DNA to affect expression of adjacent structural genes. Generally,

lac operon promoter DNA site. This is where RNA polymerase

same DNA molecule that this inactive promoter would normally signal for

lac operon operator DNA site. This is where the lactose

DNA molecule that the inactive operator would normally regulate.

cell, far from the piece of DNA that

places in the cell, or acting on different pieces of DNA. In the lac operon,

DNA sites cannot diffuse throughout the cell and only act on the adjacent

DNA. This is called acting in cis also by analogy to chemistry: in

same piece of DNA. In the lac operon, lac P and lac O

the wild-type lacZ gene because promoters only work on the DNA

chromosome on a plasmid. A plasmid is another circular DNA molecule present

E. coli

molecules of DNA and 15,000-30,000 ribosomes.

approximately 70 percent water, 15 percent protein, 1 percent DNA, 6

approximately l/500 as much DNA as is contained in a single cell of a

is also used extensively in research on recombinant DNA ("genetic

Nucleic Acids

4 Nucleic Acids

Deoxyribonucleic Acid (DNA) contains four nucleotide bases.

7.012 DNA Sequencing Section Problem

7.012 DNA Sequencing Section Problem

Given the following double-stranded piece of DNA:

MIT Biology Hypertextbook: Enzyme Mechanisms

ex. Deoxyribonuclease, or DNase catalyzes

DNA ---> dNMP nucleotides

Membrane Proteins Introduction

It must keep its molecules of life ( DNA , RNA , and its assortment of

module not yet titled

DNA recombination in phage

Culturing cells in vitro

that its DNA sequences are in order. In the event that the cellular DNA

repair its DNA, erasing as many mutations as possible, before it

proceeds into S to copy its DNA. In doing so, the cell minimizes the

their DNA has been replicated. Thus, there is an intracellular monitor

which checks on the progress of DNA replication and prevents premature

for ensuring that once a DNA molecule has been replicated in one S

DNA Structure.

DNA Structure.

DNA Structure

This is the molecular structure of DNA . A slightly more simplified diagram

DNA Structure. The four nitrogenous bases of DNA are arranged along

the sugar-phosphate backbone in a particular order (the DNA sequence),

thymine (T), while cytosine (C) pairs with guanine (G). The two DNA strands

Characteristics of Prokaryotes and Eukaryotes

They both have DNA as their genetic material.

organelles , while prokaryotes do not. The DNA of prokaryotes floats

freely around the cell; the DNA of eukaryotes is held within its

The DNA of eukaryotes is much more complex and therefore much more

extnsive than the DNA of prokaryotes.

Solutions to exercises for Mendelian Genetics Chapter

Drosophila, 0.05cM is roughly 25000 bp of DNA.

Polymerase Chain Reaction - Xeroxing DNA

Polymerase Chain Reaction - Xeroxing DNA

Polymerase Chain Reaction - Xeroxing DNA

laboratories and doctor's offices, relies on the ability of DNA-copying

produce millions of copies of a single DNA segment in a matter of hours.

In nature, most organisms copy their DNA in the same way. The PCR mimics

enzymes called polymerases make a copy of all the DNA in each

chromosome. The first step in this process is to "unzip" the two DNA

chains of the double helix. As the two strands separate, DNA polymerase

The four nucleotide bases, the building blocks of every piece of DNA,

To copy DNA, polymerase requires two other components: a supply of the

four nucleotide bases and something called a primer. DNA polymerases,

whether from humans, bacteria, or viruses, cannot copy a chain of DNA

DNA is called a primer. Once the primer is made, the polymerase can

A PCR vial contains all the necessary components for DNA duplication: a

piece of DNA, large quantities of the four nucleotides, large quantities

of the primer sequence, and DNA polymerase. The polymerase is the Taq

separates the two DNA chains in the double helix. This is done simply by

But the primers cannot bind to the DNA strands at such a high temperature, so

the primers bind or "anneal" to the ends of the DNA strands. This takes about

contains a G, it adds a C to the new chain, and so on to the end of the DNA

of a cycle, each piece of DNA in the vial has been duplicated.

DNA piece can act as a new template, so after 30 cycles, 1 million

copies of a single piece of DNA can be produced! Taking into account the

In one application of the technology, small samples of DNA, such as those

There are three related sections on DNA fingerprinting, read whichever

DNA Fingerprinting in Human Health and Society

An Interview with DNA Forensics Authority Dr. Bruce Weir

Use of DNA in Identification: A talk by Eric Lander

Southerns, Northerns, Westerns, & Cloning: Molecular Searching Techniques

These are techniques for analyzing cellular macromolecules: DNA,

Purves, Oriens, and Heller pp. 315-8. DNA-RNA hybridization is

example: the two strands of a DNA double-helix bind because they have

1) DNA-DNA. A single-stranded

DNA (ssDNA) probe molecule can form a double-stranded, base-paired

hybrid with a ssDNA target if the probe sequence is the reverse

2) DNA-RNA. A single-stranded DNA (ssDNA) probe molecule can

search for one DNA molecule, or one RNA molecule, or one protein

of different proteins. When the cell is broken open to extract DNA, RNA,

or protein, the result is a complex mixture of all the cell's DNA, RNA,

DNA cut with restriction enzymes - probed with radioactive DNA.

RNA - probed with radioactive DNA or RNA.

value - if you mix a solution of DNA with a solution of radioactive

preparing DNA, RNA and protein samples for electrophoresis.

Preparing DNA for Southern Blots

DNA is first cut with restriction enzymes and the resulting

double-stranded DNA fragments have an extended rod conformation without

different units for DNA, RNA, and protein:

DNA: Molecular weight is measured in base-pairs, or

On most gels, one well is loaded with a mixture of DNA, RNA, or

Sample 2 is what a sample of total DNA cut with a restriction enzyme,

Staining DNA

DNA is stained with ethidium bromide (EtBr), which binds to nucleic

aids. The DNA-EtBr complex fluoresces under UV light.

After the DNA, RNA, or protein has been separated by molecular

used. DNA, RNA, and protein stick well to nitrocellulose in a

The DNA, RNA, or protein can be transferred to nitrocellulose in

Note: In a Southern Blot, the DNA molecules in the gel are

probe to hybridize to them. To do this, the DNA is transferred using a

strongly alkaline buffer, which causes the DNA strands to separate -

in a blocking solution which contains a high concentration of DNA, RNA,

Radioactive DNA probes for Southerns and Northerns

double-stranded DNA fragment. The process usually begins with a

long, this results in 2 to 10 DNA fragments of different lengths. If the

identified on the gel. The band is then cut out of the gel and the DNA

the isolated DNA is a pure population of identical double-stranded DNA

The DNA restriction fragment (template) is then labeled by

1) The template DNA is denatured - the strands are separated - by

2) A mixture of DNA hexamers (6 nucleotides of ssDNA) containing all

base-pair. They pair at many sites along each strand of DNA.

3) DNA polymerase is added along with dATP, dGTP, dTTP, and radioactive

one that is incorporated into the DNA strand) is synthesized from

This process is diagrammed below (labeled DNA shown in gray):

DNA copy of both strands of the template for use as a probe.

end up with a plasmid which contains a fragment of human DNA which

of the actin gene from another organism and human chromosomal DNA.

DNA-DNA hybridization is usually used for this.

Although Southern blotting involves DNA-DNA hybridization, it is

not a useful procedure for cloning a gene. If we were to cut human DNA

the human actin gene DNA from either the gel or the filter because at

For this reason, separating the DNA fragments by molecular

separated, each containing a different fragment of human DNA.

1) Isolate genomic (chromosomal) DNA from human cells.

2) Create a plasmid library of human DNA restriction fragments. This

different plasmid with a different inserted piece of human DNA.

make their plasmid DNA single-stranded, and bind the DNA onto the

filter. There are now spots of single-stranded plasmid DNA on the

spot corresponding to a place on the filter where DNA from the human

in broth and extract their plasmid DNA. It contains a fragment of human

DNA containing the actin gene.

Solving Problems

Solving Problems

You extract genomic DNA from a red, a pink, and a white plant; digest

nitrocellulose. You have a cDNA for the R gene which you use as a

1) What is the alteration in the R-gene DNA of the mutant r allele?

EcoRI: Digestion of this region of DNA should produce 5 fragments in

-Because the probe is a cDNA, that is, it only includes the

important. Also, since hybridization occurs after the DNA fragments have

HindIII: Digestion of this region of DNA should produce 5 fragments in

the DNA at all EcoRI sites and at all HindIII sites. This should

altered between the two DNA's.

sequencing the DNA of the R and r alleles to find the exact differences.

half as much DNA in each band.

digestion. Assuming that the DNA is the same in both sexes (How could

by doing a Southern blot of DNA from males and from females, probed with

throughout - that would require sequencing the DNA from both - but it