messenger RNA (mRNA) is transcribed (produced) through reading of the codons of DNA. The mRNA produced during transcription is a single‐stranded complement to the DNA but with the base uracil instead of thymine. Subsequently, mRNA undergoes a series of posttranscriptional modifications: The introns are spliced out, a cap is added at the 5′ end of the molecule, and a string of adenylate residues (poly‐A tail) is added to the 3′ end. The mRNA is then transported out of the nucleus into the cytoplasm, where it is translated into protein by means of cellular machinery called the ribosomes. Many excellent resources describe the very complicated process of transcription and translation (e.g. Strachan and Read 1996).
Figure 2.4 The genetic code and abbreviations for amino acids.
Genes and Alleles
The physical site or location of a gene is called its locus. At any particular gene locus, there exist different forms of the gene, called alleles. These alleles are unique from each other due to variations of the nucleotides or structure in the gene, which may result in altered function of the gene and/or gene product. These alleles are analogous to the factors of inheritance and variation identified in the 1800s by Mendel. Except on the sex chromosomes of males, an individual has two alleles at each locus. In some instances, a gene may exhibit homozygosity in its two alleles, meaning that both alleles are indistinguishable from each other. Such an example is the presence of two specific disease‐causing alleles of the HBB gene that together cause sickle cell anemia. In the heterozygous state, the two alleles can be distinguished from each other. Males with a normal chromosome complement represent an exception to this convention as they are “hemizygous” for loci on the X chromosome for which they do not have a Y chromosome complement.
The difference between two alleles may be as subtle as a single base‐pair (bp) change, such as the adenine‐to‐thymine substitution in the HBB gene that alters the B chain of hemoglobin A from its wild type to its hemoglobin sickle cell state. Alternatively, allelic differences can be as extensive as large, multicodon deletions, such as those observed in Duchenne muscular dystrophy. Some bp changes have no deleterious effect on the function of the gene; nevertheless, these functionally neutral changes in the DNA still represent different forms of a gene. Rare changes in the genetic code that cause functional change of the gene or protein are often termed “mutations,” while functionally neutral changes that are more common in the population (>1%) are given the name “polymorphism.” The term “pathogenic variant” is also commonly used to refer to rare genetic changes that cause functional change in the gene or protein. “Mutation” and “pathogenic variant” may be used interchangeably throughout the chapter.
Figure 2.5 Central dogma of genetics: DNA → RNA → protein.
(Source: Reprinted by permission from Jorde et al. (1995).)
Differences in alleles can be detected via laboratory testing. The ability to detect allele differences accurately within families, between families, and between laboratories is critically important for tracking the alleles that may be involved in Mendelian and genetically complex common disorders through linkage or association analysis. Allele detection strategies may be as simple as the presence (+) or absence (−) of a deletion or point mutation or as complicated as assessing the allele size in bp of DNA. The latter application is common when highly polymorphic microsatellite repeat markers are used in linkage analysis.
Genes and Chromosomes
Genes are organized as linear structures called chromosomes, with many thousands of genes on each chromosome. Each chromosome has distinguishable sites, known as centromeres that aid in cell division and in the maintenance of chromosome integrity. The centromere is visualized as the central constriction on a chromosome, and it separates the p (short) and the q (long) arms from one another. The centromere enables correct segregation of the duplicated chromosomal material during meiosis and mitosis. Telomeres are present at both ends of the chromosome and are required for stability of the chromosomal unit.
Using appropriate staining techniques, the chromosomes in a cell can be analyzed under the microscope following cell culture and the arrest of cell division at metaphase (when the chromosomes have duplicated and condensed). At this stage of the cell cycle, a chromosome has two double‐stranded DNA molecules. Together, the strands are called sister chromatids. The sister chromatids are held together by the centromere. Photographs are magnified, and the chromosomes are arranged into a karyotype. The normal human chromosome complement consists of 46 chromosomes arranged in 23 pairs, with one member of each pair inherited from each parent (Figure 2.6). The first 22 pairs, called autosomes, are arranged according to size and are the same in males and females. The pair of sex chromosomes generally predicts an individual’s biological sex. Most females have two X chromosomes, whereas males have one X chromosome inherited from the mother and one Y chromosome inherited from the father. Therefore, the sex of an individual is primarily determined by the contribution of the father.
Figure 2.6 A G‐banded human male karyotype.
(Source: Courtesy of Mazin Qumsiyeh, Duke University Medical Center, Durham, NC.)
Because two copies of each chromosome are present in a normal somatic (body) cell, the human organism is diploid. In contrast, egg and sperm cells have haploid chromosomal complements, consisting of a single member of each chromosome pair. The correct number of chromosomes in the normal human cell was finally established in 1956, three years after the double‐helical structure of DNA was described, when Tjio and Levan (1956) demonstrated unequivocally that the chromosomal complement is 46.
Regions of chromosomes are defined by patterns of alternating light and dark regions called bands, which become apparent after a chemical treatment has been applied. One of the most common types of banding process, called Giemsa or G banding, involves digesting the chromosomes with trypsin and then staining with a Giemsa dye. G banding identifies late‐replicating, heterochromatic regions of DNA; these are the dark bands. Other chemical processes will produce different banding patterns and identify unique types of DNA.
A specific genetic locus can then be defined quite precisely along a chromosome, such as the gene FRAXA (fragile X syndrome), which is located on the X chromosome at band q27.3. Alternatively, its localization may be specified as an interval flanked by two genetic markers. Any two loci that occur on the same chromosome are considered to be syntenic or physically linked. The location of two loci on the same arm of the chromosome is specified by their positions relative to each other and to the centromere. The gene closer to the centromere is termed centromeric or proximal to the other; similarly, the gene further from the centromere is distal or telomeric to the other (Figure 2.7).
Figure 2.7 The myotonic dystrophy (DM) and insulin receptor (INSR) genes are distal (telomeric) to the ryanodine receptor 1 and CADASIL, respectively; RYR1 and CADASIL are proximal (centromeric) to DM and INSR, respectively.
The
