We already had a look at chromosomes in lecture 1 and the following terms should be familiar:
Human cells are diploid, that is they contain two of (almost) every gene. They do so by having two copies of each autosome, (chromosomes 1-22) and two sex chromosomes (either XX or XY). The normal human karyotype when viewed down the microscope at mitotic metaphase is thus either 46 XX or 46 XY. (Meaning 46 coloured blobs, two of which are XX or XY).
This picture shows a normal male mitotic metaphase spread next to an interphase nucleus.
The primary constriction is the centromere, visible in the above picture as the point where the two chromatids remain attached, but also containing the kinetochore, the point of spindle attachment. Secondary constrictions are usually only found as the stalks connecting the short arms of the two groups of acrocentric chromosomes.
When microscopes were improved to the point that the human karyotype could be reliably discerned (in the 1950s) the chromosomes could be grouped on the basis of their relative sizes and the relative lengths of their two arms, i.e. the positions of their centromeres. Now, banding techniques make it possible to identify each chromosome.
If chromosomes are treated briefly with proteinase before staining then each chromosome has a characteristic banding pattern.
The two chromosome arms are refered to as p and q (short and long respectively). Bands are numbered from the centromere. As microscopes improved the coarse banding patterns were refined by the addition of further levels of numbering so that band 9q34.1 means the 1st subband of the 4th subband of the 3rd band of the long arm of chromosome 9! Some regions of the chromosomes are uniformly staining and late replicating. They contain few (if any) genes and are composed primarily of tandemly repeated DNA sequences (satellite DNA). These are called constitutive heterochromatin. (The inactive X chromosome in female cells is also late replicating and is called facultative heterochromatin.) Centromeres, the points of attachment of the replicated chromatids to each other and to the mitotic spindle fibres are all within constitutive heterochromatin.
No, not this sort, but "Fluorescent In Situ Hybridisation".
The technique of DNA-DNA hybridisation has been discussed in lecture 5 under the heading of Southern blotting. The ability of a single stranded DNA molecule to find and bind to its complementary strand is pretty amazing. In FISH it is exploited to the utmost. 
It is possible to visualise by hybridisation, the site of a fragment of DNA of as little as 1 - 2 kb (but more usually 40 - 50 kb) at an efficiency approaching 100%. The probe molecule is labelled with a hapten such as biotin. The biotin is located with streptavidin. The streptavidin is located with antibodies. A fluorescent dye may be conjugated to the streptavidin and to the antibodies. When the spread chromosomes are illuminated by a UV lamp, a point of fluorescence can be seen where the probe / streptavidin / antibody / fluorescent dye multilayer sandwich has built up. From three to five layers of fluorescent antibodies are built up to amplify the signal.
For examples click here
Complex probes made from entire chromosomes can be made. In this way the presence that chromosome can be ascertained and also whether it has been subject to any rearrangements. This picture shows a probe made from chromosome 22 which has been used to "paint" a cell line which has a small mysterious extra chromosome. The probe hybridises to this as well as to the two normal chromosome 22s showing that the small "marker" chromosome is derived fromm chromosome 22.
If there is some reason to suspect that an embryo may have abnormal chromosomes, for instance maternal age or past history of early spontaneous abortions, it is usual to check.
At about 12 - 14 weeks of gestation it is possible to obtain foetal cells by amniocentesis. In this technique a needle is inserted into the amniotic sac and fluid is drawn off. Foetal cells, amniocytes, are suspended in the fluid and these can be cultured and their metaphase chromosomes examined. One of the commonest chromosome abnormalities is Down syndrome, the figure links to the relevant passage later in this lecture.
If a paint is made from a human chromosome it can be applied to the chromosomes of other species. This image is of marmoset chromosomes to which a human chromosome 8 paint has been applied. This shows homology with the short arm of marmoset chromosome 13, and the whole of marmoset chromosome 16. Sherlock et al., (1996) Homologies between human and marmoset (callithrix jacchus) chromosomes revealed by comparative chromosome painting. Genomics 33: 214-9
One of the commonest mutations is a change in the chromosome number but it is also one of the most damaging occurences. Very few mutations which cause visible changes in the autosomes are compatible with life. Conor and Ferguson-Smith make the analogy that if the length of the human haploid genome was drawn stretching from London to New York, the smallest visible deletion (about 4Mb) would represent about an 8 km gap and that on this scale, the average gene would be about 30 m long. So even the smallest gap will usually contain many genes. About 20% of conceptions have some sort of chromosomal disorder but because of the lethal effects of such disorders, the number actually born is only about 0.6%.
| Defect | frequency |
| triploidy | 10% |
| tetraploidy | 5% |
| trisomy | 30% |
| Turner syndrome (45 X) | 10% |
| other | 5% |
| Total | 60% |
Euploidy is the category of chromosome changes which involve the addition or loss of complete sets of chromosomes.
The possession of one complete extra set of chromosomes is usually caused by polyspermy, the fertilisation of an egg by more than one sperm. Such embryos will usually spontaneously abort.
This is usually the result of a failure of the first zygotic division. It is also lethal to the embryo. Any other cell division may also fail to complete properly and in consequence a very small proportion of tetraploid cells can sometimes be found in normal individuals.
Aneuploidy is the category of chromosome changes which do not involve whole sets. It is usually the consequence of a failure of a single chromosome (or bivalent) to complete division.
All autosomal monosomies are lethal in very early embryogenesis. They do not even feature in the table above because they abort too early even to be recognised as a conception.
The incidence of trisomy 21 rises sharply with increasing maternal age.
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Most cases arise from non disjunction in the first meiotic division, the father contributing the extra chromosome in 15% of cases. A small proportion of cases are mosaic and these probably arise from a non disjunction event in an early zygotic division. About 4% of cases arise by inheritance of a translocation chromosome from a parent who is a balanced carrier. The symptoms include characteristic facial dysmorphologies, and an IQ of less than 50. Down syndrome is responsible for about 1/3 of all cases of moderate to severe mental handicap.
The incidence is about 1 in 5000 live births. 50% of these babies die within the first month and very few survive beyond the first year. There are multiple dysmorphic features. Most cases, as in Down's syndrome, involve maternal non-disjunction. Again, a significant fraction have a parent who is a translocation carrier.
Incidence ~1 in 3000. Again most babies die in the first year and many within the first month.
Because of X inactivation and because of the paucity of genes on the Y chromosome, aneuploidies involving the sex chromosomes are far more common than those involving autosomes.
The incidence is about 1 in 500 female births but this is only the tip of the iceberg, 99% of Turner syndrome embryos are spontaneously aborted. Individuals are very short, they are usually infertile. Characteristic body shape changes include a broad chest with widely spaced nipples and may include a webbed neck. IQ and lifespan are unaffected.
The incidence at birth is about 1 in 1000 males. Testes are small and fail to produce normal levels of testosterone which leads to breast growth (gynaecomastia) in about 40% of cases and to poorly developed secondary sexual characteristics. There is no spermatogenesis. These males are taller and thinner than average and may have a slight reduction in IQ. Very rarely more extreme forms of Kleinfelter's syndrome occur where the patient has 48, XXXY or even 49, XXXXY karyotype. These individuals are generally severely retarded.
Incidence 1 in 1000 male births. May be without any symptoms. Males are tall but normally proportioned. 10 - 15 points reduction in IQ compared to sibs? More common in high security institutions than chance would suggest? Strangely, although they are fertile they do not seem to transmit the either this condition or Kleinfelter's syndrome.
About one woman in 1000 has an extra X chromosome. It seems to do little harm, individuals are fertile and do not transmit the extra chromosome. They do have a reduction in IQ comparable to that of Kleinfelter's males.
From time to time a cell sustains damage from for instance an energetic cosmic ray particle passing through and leaving behind a trail of ionisation. This may lead to chromosome breakage. The repair systems in the nucleus will do their best to make good the damage and if this involves only one break they may be able to do so with no errors. However, if more than one break has occured they may become rejoined in the wrong combinations. This can lead to one of a spectrum of possibilities.
Rearrangements where there is no visible loss or gain of genetic material are balanced. They include:
In an inversion, a piece of chromosome is lifted out, turned arround and reinserted. If this includes the centromere then the inversion is termed pericentric. If it excludes the centromere then it is a paracentric inversion. The two have slightly different genetic consequences. 1% of the UK population are heterozygous for a pericentric inversion of chromosome 9. This is absolutely without genetic consequences.
At meiosis hetrozygous inverted chromosomes have difficulty pairing and can only do so by the formation of a loop. If a recombination event occurs within the inverted loop the consequence will be a duplication and a deletion, see the gametes drawn at the bottom of the figure. If the inversion is paracentric then the centromere itself may be duplicated (which gives rise to a dicentric fragment which will try to go to both poles at anaphase I with dire consequences) or deleted (giving rise to an acentric fragment which gets left behind on the metaphase plate).
In a balanced translocation there is no net gain or loss of chromosomal material, two chromosomes have been broken and rejoined in the wrong combination. The figure shows a translocation between the imaginary chromosomes "M" and "N". Balanced reciprocal translocation is unlikely to have any severe consequence for the cell because, even if one of the breakpoints lies within a gene, most mutations are recessive (lecture 6).
It is possible that an oncogene may be activated by the translocation and this can lead to cancer. The classic example is the chromosome 9 / chromosome 22 reciprocal translocation in chronic myeloid leukaemia. (Discussed in the final lecture).
Translocations can however give rise to difficulties with reproduction. During the first meiotic prophase, the chromosomes align in pairs to form bivalents. However, a heterozygote for a reciprocal translocation forms instead a 'tetravalent'. 
The chromosomes will segregate in the first meiotic division. Many possibiliites are open, only one, the one shown in the figure, leads to balanced gametes. This is known as "alternate segregation" where the two intact chromosomes must both move to one pole and the two translocation chromosomes must both move to the other.
Deletions may either be either interstitial or terminal. If big enough to be visible a deletion must be removing many genes and will probably give rise to a severe phenotype. See above. An example of an interstitial deletion is the 15q- deletion which causes either Angelman or Prader-Wili syndromes. Many independent deletions in this region do indeed remove many genes including the two responsible for the two syndromes. You will find many more examples, try for instance searching OMIM with the acronym WAGR (which stands for Wilms tumour, Aniridia, ambiguous Genitalia and mental Retardation). See also the table on page 129 of Connor and Ferguson-Smith
Terminal deletions have only one breakpoint, they extend to the telomere. For example the karyotype shown on the right is of a baby with Cri du chat syndrome in which a small part of the distal region of the short arm of chromosome 5 is deleted. Babies with this syndrome have a combination of symptoms which include pinched facial features, mental retardation and developmental delay. The characteristic feature, for which the syndrome is named is a "mewing" cry. The charateristic cry may be separated genetically from the facial dysmorphology and developmental delay, since a small terminal deletion may have the cry only whereas a larger deletion extending further towards the centromere will include the genes whose hemizygosity is responsible for the other syptoms.
An unbalanced translocation may arise spontaneously and is also likely to arise as an offspring of a balanced carrier. There are likely to be symptoms which may be severe. Their exact nature will be unpredictable. Such translocation chromosomes are extremely useful to science in helping to pinpoint genes resposible for the conditions expressed by their bearers.
Robertsonian translocations are a special case of 'almost balanced' translocations. Robertsonian translocations involve any two out of chromosomes 13, 14, 15, 21 and 22. These chromosomes are all acrocentric, that is, the centromere is very close to one end. The short arms contain few, if any, genes except for many tandemly repeated copies of the ribosomal RNA genes. Every diploid cell thus contains 10 copies of the block of repeated genes. A Robertsonian translocation is a fusion between the centromeres of two of these chromosomes with loss of the short arms forming a chromosome with two long arms, one derived from each chromosome. 
The loss of the short arms does not matter, each cell still has eight copies of the rRNA gene block and that, apparently, is enough. In the same way as balanced reciprocal translocation carriers have difficulties at meiosis in ensuring the correct segregation of the chromosomes to make a balanced set, so too do Robertsonian translocation carriers. In their case the chromosomes pair in meiotic prophase to form a trivalent and balanced gametes will only be formed when the translocation chromosome goes to the opposite pole to both of the normal chromosomes. Robersonian carriers therefore suffer a similar reduction to their fertility as do carriers of reciprocal translocations and couples in which one of the partners has a translocation may have a number of early spontaneous abortions. Robertsonian translocations involving chromosome 21 possess a special problem of their own, one of the possible unbalanced gametes will contain effectively two copies of chromosome 21 (when the translocation chromosome and chromosome 21 segregate to the same pole). They are thus at risk of producing a baby with Down syndrome.
A chromosome can split "the wrong way" in mitosis (or meiosis II) so that both long arms remain attached and move to one pole, and both short arms do likewise moving to the other pole. The consequence is the formation of an isochromosome. These are simultaneously duplicated for the genes in the retained arm and deleted for the genes in the other. The prognosis is poor except for iXq (isochromosome of the long arm of the X).
A mutation event which removes both telomeres can be repaired by sealing the ends together forming a ring chromosome. This will be deleted for genes at both ends of the chromosome. the symtoms will depend on the extent of the deletion. Surprisingly, ring chromosomes are mitotically stable. One might expect them to get hopelessly entangled during DNA replication and at the very least to be concatenated when the time comes to seperate to the two poles. While this undoubtedly happens, it is not a frequent occurence.
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