What is DNA Fingerprinting

What is DNA Fingerprinting?

The chemical structure of everyone's DNA is the same. The only difference between people (or any animal) is the order of the base pairs. There are so many millions of base pairs in each person's DNA that every person has a different sequence.

Using these sequences, every person could be identified solely by the sequence of their base pairs. However, because there are so many millions of base pairs, the task would be very time-consuming. Instead, scientists are able to use a shorter method, because of repeating patterns in DNA.

These patterns do not, however, give an individual "fingerprint," but they are able to determine whether two DNA samples are from the same person, related people, or non-related people. Scientists use a small number of sequences of DNA that are known to vary among individuals a great deal, and analyze those to get a certain probability of a match.

How is DNA Fingerprinting Done?

1. Performing Sourthen Blot

The Southern Blot is one way to analyze the genetic patterns which appear in a person's DNA. Performing a Southern Blot involves:

1. Isolating the DNA in question from the rest of the cellular material in the nucleus. This can be done either chemically, by using a detergent to wash the extra material from the DNA,or mechanically, by applying a large amount of pressure in order to "squeeze out" the DNA.

2. Cutting the DNA into several pieces of different sizes. This is done using one or more restriction enzymes.

3. Sorting the DNA pieces by size. The process by which the size separation, "size fractionation," is done is called gel electrophoresis. The DNA is poured into a gel, such as agarose, and an electrical charge is applied to the gel, with the positive charge at the bottom and the negative charge at the top. Because DNA has a slightly negative charge, the pieces of DNA will be attracted towards the bottom of the gel; the smaller pieces, however, will be able to move more quickly and thus further towards the bottom than the larger pieces. The different-sized pieces of DNA will therefore be separated by size, with the smaller pieces towards the bottom and the larger pieces towards the top.

4. Denaturing the DNA, so that all of the DNA is rendered single-stranded. This can be done either by heating or chemically treating the DNA in the gel.

5. Blotting the DNA. The gel with the size-fractionated DNA is applied to a sheet of nitrocellulose paper, and then baked to permanently attach the DNA to the sheet. The Southern Blot is now ready to be analyzed.

In order to analyze a Southern Blot, a radioactive genetic probe is used in a hybridization reaction with the DNA in question (see next topics for more information). If an X-ray is taken of the Southern Blot after a radioactive probe has been allowed to bond with the denatured DNA on the paper, only the areas where the radioactive probe binds [red] will show up on the film. This allows researchers to identify, in a particular person's DNA, the occurrence and frequency of the particular genetic pattern contained in the probe.

2. Making a radioactive probe
1. Obtain some DNA polymerase [pink]. Put the DNA to be made radioactive (radiolabeled) into a tube.

2. Introduce nicks, or horizontal breaks along a strand, into the DNA you want to radiolabel. At the same time, add individual nucleotides to the nicked DNA, one of which, *C [light blue], is radioactive.

3. Add the DNA polymerase [pink] to the tube with the nicked DNA and the individual nucleotides. The DNA polymerase will become immediately attracted to the nicks in the DNA and attempt to repair the DNA, starting from the 5' end and moving toward the 3' end.

4. The DNA polymerase [pink] begins repairing the nicked DNA. It destroys all the existing bonds in front of it and places the new nucleotides, gathered from the individual nucleotides mixed in the tube, behind it. Whenever a G base is read in the lower strand, a radioactive *C [light blue] base is placed in the new strand. In this fashion, the nicked strand, as it is repaired by the DNA polymerase, is made radioactive by the inclusion of radioactive *C bases.

5. The nicked DNA is then heated, splitting the two strands of DNA apart. This creates single-stranded radioactive and non-radioactive pieces. The radioactive DNA, now called a probe [light blue], is ready for use.

3. Creating a Hybridization Reaction
1. Hybridization is the coming together, or binding, of two genetic sequences. The binding occurs because of the hydrogen bonds [pink] between base pairs. Between a A base and a T base, there are two hydrogen bonds; between a C base and a G base, there are three hydrogen bonds.

2. When making use of hybridization in the laboratory, DNA must first be denatured, usually by using heat or chemicals. Denaturing is a process by which the hydrogen bonds of the original double-stranded DNA are broken, leaving a single strand of DNA whose bases are available for hydrogen bonding.

3. Once the DNA has been denatured, a single-stranded radioactive probe [light blue] can be used to see if the denatured DNA contains a sequence similar to that on the probe. The denatured DNA is put into a plastic bag along with the probe and some saline liquid; the bag is then shaken to allow sloshing. If the probe finds a fit, it will bind to the DNA.

4. The fit of the probe to the DNA does not have to be exact. Sequences of varying homology can stick to the DNA even if the fit is poor; the poorer the fit, the fewer the hydrogen bonds between the probe [light blue] and the denatured DNA. The ability of low-homology probes to still bind to DNA can be manipulated through varying the temperature of the hybridization reaction environment, or by varying the amount of salt in the sloshing mixture.

4. VNTRs
Every strand of DNA has pieces that contain genetic information which informs an organism's development (exons) and pieces that, apparently, supply no relevant genetic information at all (introns). Although the introns may seem useless, it has been found that they contain repeated sequences of base pairs. These sequences, called Variable Number Tandem Repeats (VNTRs), can contain anywhere from twenty to one hundred base pairs.

Every human being has some VNTRs. To determine if a person has a particular VNTR, a Southern Blot is performed, and then the Southern Blot is probed, through a hybridization reaction, with a radioactive version of the VNTR in question. The pattern which results from this process is what is often referred to as a DNA fingerprint.

A given person's VNTRs come from the genetic information donated by his or her parents; he or she could have VNTRs inherited from his or her mother or father, or a combination, but never a VNTR either of his or her parents do not have. Shown below are the VNTR patterns for Mrs. Nguyen [blue], Mr. Nguyen [yellow], and their four children: D1 (the Nguyens' biological daughter), D2 (Mr. Nguyen's step-daughter, child of Mrs. Nguyen and her former husband [red]), S1 (the Nguyens' biological son), and S2 (the Nguyens' adopted son, not biologically related [his parents are light and dark green]).

Because VNTR patterns are inherited genetically, a given person's VNTR pattern is more or less unique. The more VNTR probes used to analyze a person's VNTR pattern, the more distinctive and individualized that pattern, or DNA fingerprint, will be.

Pratical Applications of DNA Fingerprinting

1. Paternity and Maternity
Because a person inherits his or her VNTRs from his or her parents, VNTR patterns can be used to establish paternity and maternity. The patterns are so specific that a parental VNTR pattern can be reconstructed even if only the children's VNTR patterns are known (the more children produced, the more reliable the reconstruction). Parent-child VNTR pattern analysis has been used to solve standard father-identification cases as well as more complicated cases of confirming legal nationality and, in instances of adoption, biological parenthood.

2. Criminal Identification and Forensics
DNA isolated from blood, hair, skin cells, or other genetic evidence left at the scene of a crime can be compared, through VNTR patterns, with the DNA of a criminal suspect to determine guilt or innocence. VNTR patterns are also useful in establishing the identity of a homicide victim, either from DNA found as evidence or from the body itself.

3. Personal Identification
The notion of using DNA fingerprints as a sort of genetic bar code to identify individuals has been discussed, but this is not likely to happen anytime in the foreseeable future. The technology required to isolate, keep on file, and then analyze millions of very specified VNTR patterns is both expensive and impractical. Social security numbers, picture ID, and other more mundane methods are much more likely to remain the prevalent ways to establish personal identification.

Problems with DNA Fingerprinting

Like nearly everything else in the scientific world, nothing about DNA fingerprinting is 100% assured. The term DNA fingerprint is, in one sense, a misnomer: it implies that, like a fingerprint, the VNTR pattern for a given person is utterly and completely unique to that person. Actually, all that a VNTR pattern can do is present a probability that the person in question is indeed the person to whom the VNTR pattern (of the child, the criminal evidence, or whatever else) belongs. Given, that probability might be 1 in 20 billion, which would indicate that the person can be reasonably matched with the DNA fingerprint; then again, that probability might only be 1 in 20, leaving a large amount of doubt regarding the specific identity of the VNTR pattern's owner.

1. Generating a High Probability
The probability of a DNA fingerprint belonging to a specific person needs to be reasonably high--especially in criminal cases, where the association helps establish a suspect's guilt or innocence. Using certain rare VNTRs or combinations of VNTRs to create the VNTR pattern increases the probability that the two DNA samples do indeed match (as opposed to look alike, but not actually come from the same person) or correlate (in the case of parents and children).

2. Problems with Determining Probability

A. Population Genetics
VNTRs, because they are results of genetic inheritance, are not distributed evenly across all of human population. A given VNTR cannot, therefore, have a stable probability of occurrence; it will vary depending on an individual's genetic background. The difference in probabilities is particularly visible across racial lines. Some VNTRs that occur very frequently among Hispanics will occur very rarely among Caucasians or African-Americans. Currently, not enough is known about the VNTR frequency distributions among ethnic groups to determine accurate probabilities for individuals within those groups; the heterogeneous genetic composition of interracial individuals, who are growing in number, presents an entirely new set of questions. Further experimentation in this area, known as population genetics, has been surrounded with and hindered by controversy, because the idea of identifying people through genetic anomalies along racial lines comes alarmingly close to the eugenics and ethnic purification movements of the recent past, and, some argue, could provide a scientific basis for racial discrimination.

B. Technical Difficulties
Errors in the hybridization and probing process must also be figured into the probability, and often the idea of error is simply not acceptable. Most people will agree that an innocent person should not be sent to jail, a guilty person allowed to walk free, or a biological mother denied her legal right to custody of her children, simply because a lab technician did not conduct an experiment accurately. When the DNA sample available is minuscule, this is an important consideration, because there is not much room for error, especially if the analysis of the DNA sample involves amplification of the sample (creating a much larger sample of genetically identical DNA from what little material is available), because if the wrong DNA is amplified (i.e. a skin cell from the lab technician) the consequences can be profoundly detrimental. Until recently, the standards for determining DNA fingerprinting matches, and for laboratory security and accuracy which would minimize error, were neither stringent nor universally codified, causing a great deal of public outcry.

What is DNA ?

1. Nucleotides are the building stones of DNA.

There are 4 different nucleotides :
• dATP : deoxyadenosine triphosphate
• dGTP : deoxyguanosine triphosphate
• dTTP : deoxythymidine triphosphate
• dCTP : deoxycytidine triphosphate
For convenience, these 4 nucleotides are called dNTP's (deoxynucleoside triphosphates). A nucleotide is made of three major parts : a nitrogen base, a sugar molecule and a triphosphate. Only the nitrogen base is different in the 4 nucleotides.

Figure: The components of nucleotides. (pdf file of this picture)

2. How do the nucleotides form a DNA chain ?

Figure: From nucleotide to DNA. (pdf file of this picture) DNA is formed by coupling the nucleotides between the phosphate group from a nucleotide (which is positioned on the 5th C-atom of the sugar molecule) with the hydroxyl on the 3rd C-atom on the sugar molecule of the previous nucleotide. To accomplish this, a diphosphate molecule is split off (and releases energy). This means that new nucleotides are always added on the 3' side of the chain.

What is DNA Sequence Alignment?

To compare two or more sequences, it is necessary to align the conserved and unconserved residues across all the sequences (identification of locations of insertions and deletions that have occurred since the divergence of a common ancestor). These residues form a pattern from which the relationship between sequences can be determined with phylogenetic programs. When the sequences are aligned, it is possible to identify locations of insertions or deletions since their divergence from their common ancestor. There are three possibilities :

• The bases match : this means that there is no change since their divergence.
• The bases mismatch : this means that there is a substitution since their divergence.
• There is a base in one sequence, no base in the other : there is an insertion or a deletion since their divergence.

Figure: The comparison of sequences. A good alignment is important for the next step : the construction of phylogenetic trees. The alignment will affect the distances between 2 different species and this will influence the inferred phylogeny. There are several programs available on the net for aligning sequences. These are all based on different mathematical models to compare two or more sequences with the most optimal score for matching bases with a minimum number of gaps inserted (because you can insert a huge amount of gaps, so every base will match an other).

Example : two sequences :

TCAGACGATTG
TCGGAGCTG

How can we get the best alignment ? There are several possibilities : 1. Reduce the number of mismatches :

TCAG-ACG-ATTG
|| | | |  | |        0 mismatches  7 matches  6 gaps
TC-GGA-GC-T-G

2. Reduce the number of gaps :

TCAGACGATTG
|| ||                5 mismatches  4 matches  2 gaps
TCGGAGCTG--

3. Reduce neither the number of gaps nor the number of mismatches :

TCAG-ACGATTG
|| | | | |           2 mismatches  6 matches  4 gaps
TC-GGA-GCTG-

4. Same as 3. but one base (or gap) moved :

TCAG-ACGATTG
|| | | | | |         1 mismatch    7 matches  4 gaps
TC-GGA-GCT-G

Which of these is now the best alignment ?? There are several alignment algorithms to choose the best alignment. Let's use a simple one in this example :

D = y + sum(w_kz_k)

with :

D = distance
y : number of mismatches
w : penalty for gaps of length k
z : number of gaps of length k

Take gap penalty for gap length 1 = 2
Take gap penalty for gap length 2 = 6 (short gaps occur more frequent than long gaps)

in 1. : 0 + {(2 x 6) + (6 x 0)} = 12
in 2. : 5 + {(2 x 0) + (6 x 1)} = 11
in 3. : 2 + {(2 x 4) + (6 x 0)} = 10
in 4. : 1 + {(2 x 4) + (6 x 0)} = 9

We choose alignment 4 because it has the minimum distance.

Figure: The alignment of sequences. This is done with Clustalw 1.74, and as you can see, the more variable areas are not optimally aligned (indicated with red boxes). Therefore it is mostly necessary to improve the alignment by hand. In this case, it is obvious to improve the alignment, but in other cases it could be more difficult to make improvements.

The Principle of DNA Sequencing

The purpose of sequencing is to determine the order of the nucleotides of a gene. For sequencing, we don't start from gDNA (like in PCR) but mostly from PCR fragments or cloned genes.

1. The sequencing reaction :
There are three major steps in a sequencing reaction (like in PCR), which are repeated for 30 or 40 cycles.


1. Denaturation at 94°C :

During the denaturation, the double strand melts open to single stranded DNA, all enzymatic reactions stop (for example : the extension from a previous cycle).


2. Annealing at 50°C :

In sequencing reactions, only one primer is used, so there is only one strand copied (in PCR : two primers are used, so two strands are copied). The primer is jiggling around, caused by the Brownian motion. Ionic bonds are constantly formed and broken between the single stranded primer and the single stranded template. The more stable bonds last a little bit longer (primers that fit exactly) and on that little piece of double stranded DNA (template and primer), the polymerase can attach and starts copying the template. Once there are a few bases built in, the ionic bond is so strong between the template and the primer, that it does not break anymore.


3. extension at 60°C :

This is the ideal working temperature for the polymerase (normally it is 72 °C, but because it has to incorporate ddNTP's which are chemically modified with a fluorescent label, the temperature is lowered so it has time to incorporate the 'strange' molecules. The primers, where there are a few bases built in, already have a stronger ionic attraction to the template than the forces breaking these attractions. Primers that are on positions with no exact match, come loose again and don't give an extension of the fragment.

The bases (complementary to the template) are coupled to the primer on the 3'side (adding dNTP's or ddNTP's from 5' to 3', reading from the template from 3' to 5' side, bases are added complementary to the template).

When a ddNTP is incorporated, the extension reaction stops because a ddNTP contains a H-atom on the 3rd carbon atom (dNTP's contain a OH-atom on that position). Since the ddNTP's are fluorescently labeled, it is possible to detect the color of the last base of this fragment on an automated sequencer.

Figure 7 : The different steps in sequencing. (pdf file of this picture)Animated picture of sequencing (344 kB) Because only one primer is used, only one strand is copied during sequencing, there is a linear increase of the number of copies of one strand of the gene. Therefore, there has to be a large amount of copies of the gene in the starting mixture for sequencing. Suppose there are 1000 copies of the wanted gene before the cycling starts, after one cycle, there will be 2000 copies : the 1000 original templates and 1000 complementary strands with each one fluorescent label on the last base, after two cycles, there will be 2000 complementary strands, three cycles will result in 3000 complementary strands and so on.

Figure 8 : The linear amplification of the gene in sequencing.
1. Separation of the molecules :
After the sequencing reactions, the mixture of strands, all of different length and all ending on a fluorescently labelled ddNTP have to be separated; This is done on an acrylamide gel, which is capable of separating a molecule of 30 bases from one of 31 bases, but also a molecule of 750 bases from one of 751 bases. All this is done with gel electrophoresis. DNA has a negative charge and migrates to the positive side. Smaller fragments migrate faster, so the DNA molecules are separated on their size.

Figure 9 : The separation of the molecules with electrophoresis.(pdf file of this picture)Animated picture of gel electrophoresis (159 kB)


1. Detection on an automated sequencer :
The fluorescently labelled fragments that migrate trough the gel, are passing a laser beam at the bottom of the gel. The laser exites the fluorescent molecule, which sends out light of a distinct color. That light is collected and focused by lenses into a spectrograph. Based on the wavelength, the spectrograph separates the light across a CCD camera (charge coupled device). Each base has its own color, so the sequencer can detect the order of the bases in the sequenced gene.

Figure 10 : The scanning and detection system on the ABI Prism 377 sequencer. (pdf file of this picture)Animated picture of scanning and detection system (182 kB)

Figure 11 : A snapshot of the detection of the molecules on the sequencer.

1. Assembling of the sequenced parts of a gene :
For publication purposes, each sequence of a gene has to be confirmed in both directions. To accomplish this, the gene has to be sequenced with forward and reverse primers. Since it is only possible to sequence a part of 750 till 800 bases in one run, a gene of, for example 1800 bases, has to be sequenced with internal primers. When all these fragments are sequenced, a computer program tries to fit the different parts together and assembles the total gene sequence.

Figure 12 : The assemblage of the gene.

More information in power point and pdf format.

Nanoparticles Home in on Brain Cancer

Call them laser-guided smart bombs for brain tumors. Researchers at the University of Michigan announced the testing of a drug delivery system that involves drug-toting nanoparticles and a guiding peptide to target cancerous cells in the brain. Their study finds that via this method more of the drug can be delivered to a tumor's general vicinity. They report their findings in the November 15 issue of Clinical Cancer Research.

The researchers used a pharmaceutical called Photofrin, which is photodynamic, meaning it is activated by a laser after it has entered the bloodstream. As its primary side effect, the drug renders patients photosensitive, and they must remain out of bright sunlight and even unshaded lamps for up to 30 days after receiving treatment. Despite this major drawback, Photofrin is used in the treatment of esophageal, bladder and skin cancers. But their novel delivery system, which relies on the intravenous delivery of 40-nanometer-wide particles to carry the drug, may actually avoid much of the photosensitivity, because less Photofrin circulates in the bloodstream thanks to a peptide called F3. A sequence of 31 amino acids broken off of the protein HMGN2 (high mobility group protein 2), F3 has the ability to penetrate cell membranes. "This peptide acts as a "zip code" in that it enables the binding of the nanoparticles only to blood vessels within the tumor and not normal blood vessels," says Alnawaz Rehemtulla, a radiologist and environmental health scientist who co-authored the study. F3 can detect the expression of a protein called nucleolin, which is a marker on the surface of tumor cells.

Another problem the researchers avoided was having to deliver their medicine in such a way that it could cross the blood-brain barrier, which keeps many substances from entering the brain from the bloodstream. Typical chemotherapies must penetrate this shield to treat tumors. In this case, however, the nontoxic polyacrylamide particles didn't have to cross over via the bloodstream. "The nanoparticles do not need to cross the blood-brain barrier as they were specifically designed to target the blood vessel cells within the tumor," explains radiologist Brian Ross, one of the study's authors. "The treatment should be thought of as an antivascular treatment thereby shutting off the tumor blood flow resulting in the death of the tumor cells through starvation of oxygen and energy sources."

To test the delivery method, researchers divided 34 rats--all who received injections of cancerous cells into their brains--into different groups. Those that received no treatment or got only the laser fared poorly, dying on average within 8.5 days. Those that got Photofrin either intravenously or encapsulated in nanoparticles had a median survival time of 13 days. The group that got F3 with the Photofrin-carrying nanoparticles came through the best: they lived for, on average, 33 days; three of the five in this grouping lived for 60 days, and two of those three appeared tumor-free after six months. By using iron oxide as a contrast agent--to more easily detect where the nanoparticles ended up via MRI--the group determined that twice as much drug with the F3 peptide attached reached the tumor site--10 percent of the total amount administered--compared with when nontargeted nanoparticles were injected.

Ross says that based on the success of the study, the team is investigating if this delivery technology will work for nonphotodynamic therapies. Rehemtulla adds that if other FDA-approved chemotherapeutic agents reach their targets as successfully as Photofrin did, "then we will have developed a way to make cancer drugs more 'tumor-specific,' because they will only get into tumor vasculature and not normal vasculature. This will spare patients from normal tissue toxicity that is commonly associated with almost all chemotherapy." --Nikhil Swaminathan

British soldiers kill white rhino

British troops training in northern Kenya have shot dead a white rhinoceros after it charged at them.

The four soldiers were confronted by the adult male after they got lost at night on an exercise in the bush.

The shooting happened on Friday evening in Laikipia, north of the capital Nairobi. The area is popular for wildlife viewing.

White rhinos are killed by poachers who want their horns for use in traditional Chinese medicine or as ornaments.

Laikipia conservancy senior game warden Dickson Too said the soldiers had been "forced to shoot at it".

"We don't consider it a deliberate act of killing, they were just acting in self-defence," he said.

Mr Too added the rhino had been found the following day and the Kenya Wildlife Service had removed its horn.

Kenya Wildlife Service spokeswoman Connie Maina said British troops had a base near Laikipia and regularly underwent training in the area.

Inventor of a DNA Sequencing Technique Is Disputed

A small biotechnology company has emerged to claim that it invented a seminal technique crucial to biotechnology research. And the government says it will consider, nearly a quarter-century after the invention was made, whether it awarded the patent to the wrong party.

The United States Patent and Trademark Office has started a proceeding to determine the rightful inventor of the technique, automated DNA sequencing: scientists at the California Institute of Technology, who hold the patent, or those at Enzo Biochem, the small company.

If Enzo were to win the patent rights it could mean significant revenue for the company and could hurt Applied Biosystems, which licenses the patent in question from Caltech and dominates the DNA sequencing business. Applied Biosystems machines were the main ones used in the Human Genome Project to determine man’s genetic blueprint.

Applied Biosystems, a unit of Applera, recorded $540 million in sales of DNA sequencing machines and chemicals in the fiscal year ended June 30, accounting for 29 percent of its revenue. Caltech is estimated to have earned tens of millions of dollars from that and related patents.

Executives at Enzo, which is based in New York, say the company filed for a patent in June 1982, a few months before the Caltech scientists said they conceived of their invention. But Enzo’s application was continually rejected, delayed and amended in the patent office and remained unknown until now.

“We had to fight and fight with them,” said Eugene C. Rzucidlo, a New York patent lawyer who represents Enzo. “It’s only now that the patent office granted our claims and set up a proceeding to find who the real first inventor is.”

Patent law was changed in the 1990s to eliminate such “submarine patents,” which can lead to infringement by companies that develop a product on their own, never knowing about a patent that suddenly surfaces. But Enzo’s application is old enough to fall under the old rules.

If it were to get a patent it would last for 17 years from the date it is issued. Caltech’s patent was issued in 1998 and expires in 2015.

Spokeswomen for Caltech and Applied Biosystems said their organizations did not know enough yet to comment. But Edward R. Reines, a Silicon Valley patent lawyer who has represented Applied Biosystems, accused Enzo of trying to mine the patent system for money.

“Enzo appears to be attempting to claim credit for the invention of modern DNA sequencing 25 years after the fact when they have not brought a meaningful DNA sequencing product to market,” said Mr. Reines, who is with Weil, Gotshal & Manges.

Enzo executives disputed that, saying the company, founded in 1976, had long sold reagents for use in genetic analysis, though not sequencing machines. Enzo reported a net loss of $15.7 million in its last fiscal year on revenue of $39.8 million.

The Caltech sequencer attached a different color of fluorescent dye to each of the four chemical units of DNA, allowing the DNA sequence to be read by a machine. It made sequencing faster and set the stage for the Human Genome Project. The inventors include Leroy E. Hood, a biologist, and Michael W. Hunkapiller, who later ran Applied Biosystems.

The Caltech patents have withstood challenges before. A whistleblower lawsuit filed by a competitor of Applied Biosystems, said federal funds had been used in the invention, entitling the government to certain discounts and other rights. The government declined to pursue the lawsuit. And a former Caltech scientist filed a lawsuit claiming to be one of the inventors but lost in court.

R. Danny Huntington, a Washington lawyer who specializes in interference proceedings, said there were only about 100 such cases a year and they could take one or two years to resolve. The party with the earlier patent application date wins about two times out of three, he said.

While that would seem to favor Enzo, the patent office declaration of the interference criticizes Enzo’s patent claims for being unusually numerous, “erratically numbered” and “extraordinary in their flagrant disregard” of certain rules. The application is still not public, but lawyers said it contained about 1,200 claims.

Disease Detectives

Anytown, U.S.A., has a serious problem. One of its residents is very sick. Doctors suspect avian influenza. The disease, also called bird flu, can be devastating.

"If we do nothing," says Taylor Jones, the freckle-faced mayor of Anytown, "most likely, 70 percent of people in this town will die."

In a lab at the National Institutes of Health, student scientists Jack Grundy (left) and Erin Edwards tackle a make-believe avian-flu epidemic at this year's Discovery Channel Young Scientist Challenge.

While Jones and an epidemiologist use computer models to assess the town's risk, a virologist scans mucus samples to prepare a diagnosis. The patient, a 33-year-old named Joe Plastic, lies in a hospital isolation unit. He's struggling to breathe.

"He's starting to die," says Dr. Jayne Thompson.

The virologist, Kushal Naik, has more bad news.

"Joe is positive for avian flu, but that's not the worst part," Naik says. "We have nine specimens from other hospitals that are also positive. It's spreading."

Jayne Thompson and William Pete take a mucus sample from Joe Plastic's nose.

This crisis ends quickly, however, mainly because it's fictional. The team, ranging in age from 11 to 15, is tackling one of six 90-minute challenges at this year's Discovery Channel Young Scientist Challenge (DCYSC).

Each fall, DCYSC brings 40 middle school science fair champs to Washington, D.C., to compete for more than $100,000 in scholarships, prizes, and the honor of being named "America's Top Young Scientist of the Year." Winners must combine problem solving with quick thinking, teamwork, and the ability to explain complicated ideas clearly.

Gut navigation

This year's team competition, which had a medical theme, took place at the National Institutes of Health (NIH) in Bethesda, Md. Most challenges involved real-world medical problems. And cutting-edge NIH researchers were there to help.

"We try to deal with issues in the news," says Steve "Jake" Jacobs, head DCYSC judge. "NIH provided us with an opportunity available nowhere else on the planet."

NIH radiologist Ronald Summers explains the basics of reading a computerized tomography scan.

NIH researcher Ronald Summers, for example, studies virtual colonoscopy, a new way to screen for cancer of the colon (or large intestine). The technique combines X-ray–like computerized tomography (CT) scans with computer software to create three-dimensional videos of the inside of the colon. Doctors can then check the images for polyps, mushroomlike growths that can become cancerous.

The new diagnostic method is more comfortable for patients than the standard procedure. In that procedure, "you insert the scope into the patient's bottom and thread it through," Summers says. "A light and digital camera show you everything."

To compare the standard and new methods, students tried out each one. To perform a mock CT exam, they navigated through virtual images of five colons to spot the polyps in each. For the standard method, students threaded a 63-inch-long scope through a plastic model of a human colon. A screen displayed what was inside.

Steering the probe through the twists and folds of the colon was difficult. "I have no idea what I'm looking at," Otana Jakpor, 12, admitted at one point. Teammate Jack Grundy, 13, punctured the fake patient's intestinal wall by mistake.

Nolan Kamitaki and Anthony Hennig separate zebrafish embryos in a petri dish in the NIH labs.

Before the challenge ended, the colon explorers regrouped with teammates who had been injecting glowing proteins into see-through fish embryos. Together, the team needed to make a 3-minute, kid-to-kid video about new ways to look inside organisms.

Lunchtime

Downstairs, a different group of finalists battled another public health crisis: obesity (see "Packing Fat").

First, the team had to assemble a 500-calorie lunch from a selection of foods whose nutritional labels were hidden. The team picked a chicken wrap, a banana, carrot sticks, Fig Newtons, and milk.

The students were dismayed to learn that they'd overshot their mark: The lunch they'd assembled packed a walloping 885 calories.

Jayleen McAlpine demonstrates on a treadmill how much effort it takes to burn calories while her teammates look on.

Next, they used a chart, a treadmill, and their mathematics skills to figure out how much exercise it would take for a 125-pound person to burn off such a lunch.

After arguing about who would actually do so much exercise, they settled on four choices: an hour of basketball, an hour of tennis, 30 minutes of walking, and 30 minutes of lawn mowing.

Finally, the team created a podcast about energy balance and weight control.

"If people realized they had to do all that [exercise to burn off the calories in] a cookie, they might change their minds," Joseph Church, 14, said.

Collin McAliley, 13, was unconvinced. "It's such a good cookie, though," he said.

Grand prize

DCYSC involved more than challenges, dinners, meeting people, and having fun. On the final morning, the finalists visited an elementary school in Washington, D.C. They fielded questions, demonstrated science experiments, and helped kids with their science projects.

DCYSC competitor Joel Tinker demonstrates an experiment to two students at a Washington, D.C., school.

At the awards ceremony, the grand prize, a $20,000 scholarship, went to Nolan Kamitaki, 14, of Waiakea Intermediate School in Hilo, Hawaii.

Jacob "Pi" Hurwitz, 14, of Robert Frost Middle School in Rockville, Md., received a $10,000 scholarship. His nickname reflects his ability to recite 320 decimal digits of the number pi.

Amy David, 15, of Pinedale Middle School in Wyo., won third place and a $5,000 scholarship.

"One reason we're happy to have such bright, energetic people getting into science is that you are the next generation of leaders," NIH's Anthony Fauci told the finalists. "You are choosing a life of discovery and a probing of the unknown. It's a most unusual and extraordinary life."

Sharp Eye on the Sun

The sun is hotter than anything you can probably imagine, but that may not be the most striking thing about our closest star. The real surprise is that the sun's thin outer atmosphere, or corona, is much, much hotter than the sun's surface.

That's like the air high above a flame being hotter than the flame itself. The temperature should fall as you move away from a heat source.

A new spacecraft called Hinode has just started collecting data that might help explain this solar oddity.

The Earth-orbiting Hinode spacecraft, shown in this illustration, has telescopes and instruments for studying the sun.

cently launched through a collaboration involving Japan, Great Britain, and the United States, Hinode can collect two kinds of information about the sun. With a half-meter-wide visible-light telescope, it takes pictures of the sun's surface. It's the largest solar telescope that has ever flown into space.

Hinode's visible-light telescope shows details of the sun's turbulent surface, where great plumes of hot gas rise and fall to give the surface a speckled appearance.

Hinode also carries an X-ray telescope that detects hot gases in the sun's corona.

Hinode's X-ray telescope can record emissions that range between about 1 million and 4 million kelvins (273.15 kelvins equals 0°C or 32°F). This is an unusually wide temperature range for a detector, and it gives Hinode the power to sense the corona's calm, quiet features as well as its hot, explosive ones. Until now, scientists have been unable to study the corona in such detail.

New X-ray images of the sun reveal features known as X-ray bright points. Two examples are visible in the box.

The portrait (shown above) taken by Hinode's X-ray telescope on Oct. 28 shows features called X-ray bright points. These features, it appears, are magnetic loops that trap hot gas.

By monitoring X-ray bright points, scientists hope to better understand how the sun's corona becomes so hot. They should also get a clearer picture of how magnetic fields affect the corona.

Hinode, which means "sunrise" in Japanese, is still undergoing tests. In December, the spacecraft will officially begin a 3-year mission to unravel the sun's secrets.