KARYOTYPE ANALYSIS

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Genetic counseling is a relatively new field that allows parents the opportunity to determine the
genetic odds that their offspring may carry a particular negative (or lethal) trait. At first the counseling process was restricted to interviews, which helped create pedigree analysis charts that revealed the expression of the trait in question among various family members. As technology improved, a process called karyotype analysis was developed. This procedure provides information regarding the entire chromosomal complement of an individual, as viewed during the process of mitosis. Karyotypes were first performed using blood samples taken from youngsters and adults. Later, a prebirth process called amniocentesis was developed which allowed medical researchers to obtain samples of cells from extraembryonic fluid located in the amniotic sac, which surrounds an unborn child. Still later a more refined version of the procedure, called chorionic villi sampling (CVS), was developed that could be used earlier in the pregnancy than amniocentesis. CVS takes samples of extraembryonic cells from extensions of the chorionic membrane called villi. Karyotypes may be performed by taking (1) a blood, bone marrow, or skin sample from an adult; or (2) a sample of amniotic fluid (which contains stray cells) or extra-embryonic cells (from the chorionic villi) from an unborn child. Once the cells have been obtained for a karyotype, they are mixed with plant-derived chemicals called

lectins that stimulate mitosis.

After the required quantity of cells has been acquired, the cells are treated with a drug called

colchicine to stop the mitotic process at metaphase. Colchicine arrests the action of spindle fiber microtubules; hence ceasing the mitotic process.
The cells are placed in a hypotonic solution (see: osmosis and diffusion lab) that causes them to take on water, thereby increasing their overall size. This provides room for chromosomes to be spread out. The cells are fixed to a microscope slide,

stained and then photographed.

After they are photographed, the photo is enlarged and the chromosomes are cut out and matched up as pairs. Chromosomes may be distinguished from one another based upon several key characteristics: (1) the length of the arms of the chromosome; a pattern that is established by the position of the centromere along the arms, (2) shape and (3) general appearance of the chromosome, such as size and placement of bands. Karyotype analysis provides a mechanism to determine if nondisjunctional diseases, such as Down syndrome (Trisomy 21), are present in the fetus.

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It also allows for detection of other nondisjunctional diseases, such as Klinefelters syndrome (XXY) or Turners syndrome (XO), which may otherwise go unnoticed until puberty (or even later). It also provides a look at the chromosomes to see if there are any missing segments (deletions) or translocations that may have occurred during crossing-over. Refer to your biology text to help review normal and abnormal karyotype charts. As technology advanced, more sophisticated versions of karyotyping emerged. The Human Genome Project (HGP) is an ongoing worldwide effort to use new biotechnological advances in the field of molecular biology (genetic engineering, recombinant DNA technology, etc.) to map all nucleotides grouped as genes in human chromosomes. The human genome is contained in the 23 pairs of chromosomes located in the nucleus of each somatic cell. The project was originally coordinated by the National Institutes of Health (NIH) and the Department of Energy (DOE). The director of the program was Dr. James Watson (of Watson and Crick). The HGP used new techniques such as DNA extraction, gel electrophoresis, polymerase chain reaction (PCR) and many others to accomplish much more than the karyotype procedure could. The HGP has located just about every genes position on each of the human (46) chromosomes. The project began in 1988 and is now, for all intents and purposes, completed. Chromosome #22 was completely mapped by February 2000. It was the first human chromosome to be completed. Approximately 100,000 gene segments are carried in our 23 pairs of chromosomes and the rest of the DNA sequences are called fillers, spacers, RFLPs, or nonsense codes (junk DNA). Research continues to attempt to determine the value of the non-gene nucleotide sequences in the human genome. There are so many nucleotides within a single set of 46 chromosomes (23 pairs) that scientists doing karyotype/DNA analysis work always use the haploid cells (gametes) of the body whenever possible. This restricts them to only a single set of 23 chromosomes, which contain +/- 3,000,000,000 (3 x 109) nucleotides. Amazingly enough, as little as 5% of our total DNA complement is used as genes. Recall from lecture class that each chromosome contains the same linear sequences of DNA, (except for X/Y and mutations). Mapping of the human genome will yield many benefits, especially in the field of health. Samples of cells from unborn children will be taken and their analysis will reveal more than what the old karyotype test did. That test merely showed if the cell had a full set of properly structured chromosomes. It was limited to revealing only a few types of genetic problems, such as those caused by nondisjunctions. The new, extended form of karyotype analysis will also be able to tell the researcher if the child has gene mutations or if any sort of disease genes are present. The maps of the human genome are of two varieties and provide these two types of information (1) genetic maps = location of genes and their patterns of inheritance and (2) physical maps, such as restriction maps, which describe the structural characteristics of the DNA molecule itself. This is the outcome of the work that began in the classical and chromosomal eras of genetics!

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Remember Thomas Hunt Morgan? He did gene linkage maps of Drosophila in the early 1900s.

Mendels traits and factors became Morgans genes.

What about Griffith and bacterial transformation? That is where all this began! See? Research builds a stockpile of information that technology drives to varied uses! Your instructor will provide you with sample karyotype charts to analyze. You will review the karyotypes provided, cut out the chromosomes and create a final chart showing the matching chromosome pairs. Your completed charts will allow you to predict whether the individual in question is male or female and also whether their number of chromosomes is correct or incorrect.

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