THE GENETICAL SOCIETY OF GREAT BRITAIN ABSTRACTS of Papers presented at the HUNDRED AND EIGHTY-SEVENTH MEETING of the Society held on 5th, 6th and 7th JULY 1978 at the University of .

1. SATELLITE AND REPETITIVE DNA IN EUKARYOTIC CHROMOSOMES THE ISOLATION OF HUMAN Y CHROMOSOME SPECIFIC GENE SEQUENCES A.-M. SALMASI, S. MALCOLM and R. WILLIAMSON Department of Biochemistry, St. Mary's Hospital Medical School, Praed Street, London W2 IPG

HumanY chromosome-enriched gene sequences were isolated by hybridising radioactive (nick-translated) male DNA isolated from a normal male placenta to 10,000-fold excess of cold (non-labelled) female DNA isolated from a placenta of a normal female foetus. The hybridisation process was run to Cot l0, at which the non-hybridised material (15-20 per Cent of the input) was isolated by HAP chromatography analysis and rehybridised to a further 104-fold excess of non-labelled female DNA to a Cot of l0. The single-stranded material (50-60 per cent) was isolated at the end of the reaction and hybridised again to lO'-fold excess of cold female DNA to Cot I 0. The non-hybridised material (60 per cent) was found to be about 1 per cent of the original starting input (male-enriched gene sequences) - Thereannealing profiles of this probe to cold DNA of both male and female were compared. This revealed the existence of male-specific mid-repetitive and low-repetitive gene sequences shown by the existence of a difference between all the points of the reannealing curves. In situ hybridisation of the male-enriched sequences to a normal male chromosome preparation showed a high affinity of hybridisation to the distal segment of the long arm of theY chromo- some as compared to both the Y chromosome and the other groups of chromosomes.

LOCALISATIONOF 5S RIBOSONIAL RNA TO lq 41-43 BY IN SITU HYBRI- DISATION IN A FAMILY WITH AN INTERSTITIAL TRANSLOCATION OF THE LONG ARM OF CHROMOSOME I S. JANE FENNELL, SUSAN MALCOLM, R. WILLIAMSON and M. A. FERGUSONSMITH* Department of Biochemistry, St. Mary's Hospital Medical School, University of London, London, and *Department of Medical , Royal Hospital for Sick Children, Glasgow The5S ribosomal gene complement of a family with an interstitial translocation on the long arm of chromosome 1 close to the 5S ribosomal RNA locus was studied in situ hybridi- sation techniques. The deletion, shown by banding studies to occur between lq 25-32, did not include the 5S genes. Mouse 5S ribosomal RNA was labelled with 1251tohigh specific activity and hybridised to chromosomal DNA. This confirms the assignment of ribosomal 5S genes to lq, 41-43 which is in agreement with Steffensen, Chu, Speert, Wall Meilinger and Kelch (Human Genetics, 36, 25, 1977).

REPEATEDNUCLEOTIDE SEQUENCE CHANGES THAT ACCOMPANY SPECIATION IN THE TRITICACEAE RICHARD FLAVELL, MICHAEL O'DELL and DEREK SMITH Cytagenetics Department, Plant Breeding Institute, Trumpington, Cambridge CB2 2LQ

Speciationis accompanied by changes in the repeated nucleotide sequence complements of higher plant and animal chromosomes. New families of repeated sequences are often 413 414 GENETICAL SOCIETY OF GREAT BRITAIN

created. Changes in the number of copies of sequences in existing families also occur. These conclusions will be illustrated by the repeated sequence differences in wheat, oats, barley and rye, which have diverged from a common ancestor. Common bread wheat is an allo- hexaploid (2n =6x=42),containing one diploid from the genus Triticum and probably two diploid from the genus Aegilops. Some of the most highly repeated sequences in Aegilops species are not amplified in Triticunz species while others are amplified to a lesser extent. Thus the diploid parents of hexaploid wheat can be distinguished from one another by techniques which assay families of highly repeated sequences.

HIGHLYREPEATED DNA SEQUENCES IN PLANT GENOMES: A SATELLITE DNA OF CEREALS W. L. GERLACH*, E. S. DENNIS and W. J. PEACOCK CSIRO Division of Plant Industry, P.O. Box 1600, Canberro City, Australia 2601. *Present Address: Plont Breeding Institute, Mans Lone, Trumpington, Cambridge CB2 2LQ

Asatellite DNA from cereal plants has been eharacterised to see whether the rules which have emerged for this class of DNA in Drosophila and otber animals (viz distinctive chromo- somal arrangements both in position and amount, conservation of satellite sequence etc.) apply to the plant kingdom. The cereals have been chosen because they are amongst the cytogenetically best characterised plants. The satellite DNA has been isolated from hexaploid wheat, Triticum aestivuns var. Chinese Spring, and barley, Herdeum vulgare var. Clipper using Ag— Cs5SO4 buoyant density gradients.It is AT-rich relative to mainband DNA and consists of a highly repeated sequence with limited heterogeneity. Both physical properties and biochemical characterisation indicate that the sequence of the satellite has been highly conserved relative to mainband DNA during of the cereals. Using in situ hybri- disation a large proportion of the satellite has been located in blocks on a number of wheat chromosomes, including all seven B genome chromosomes. Thus, this satellite is comparable to the satellites of Drosophila and other animals in showing segmental chromosomal specificity.

REPETITIVESEQUENCES AND NON-CODING DNA P.M.B. WALKER MRC Mammalian Genome Unit, Edinburgh

Therecent history of research in eukaryotie DNA has been one of the discovery of an ever-increasing list of kinds of sequences which share two properties: they do not code for proteins and they have, at present, no known function. They include satellites and spacers, the sequences at the 5' and 3' termini of messenger RNA and leader and insertion sequences. They probably also include palindromes, polypyrimidine tracts, many of the sequences which specify HnRNA and the middle-repetitive DNA sequences. Are they all related, do they have to have a function, or could they be the result of the vagaries of sequence trans- location, duplication and crossing-over, which the genome can accommodate, as is perhaps shown by the many instances in which two closely related species maintain very different C-values of their DNA?

THEARRANGEMENT OF SATELLITE DNA SEQUENCES ON INDIVIDUAL HUMAN CHROMOSOMES CHRISTOPHER J. BOSTOCK MRC Mammalian Genome Unit, King's Buildings, West Mains Rood, Edinburgh

Humansatellite III DNA can be purified by a combination of Cs2SO4+Ag and CsCl density gradient centrifugations to give an apparently homogeneous density component. Like many other satellite DNAs, human satellite III reassociates rapidly, indicating that it is composed of a sequence of low complexity. However, analysis of the DNA fragments GENETICAL SOCIETY OF GREAT BRITAIN 415 produced by digestion with either Hae III or Eco Ri restriction endonucleases shows that human satellite III is composed of at least three distinguishable sequences, and that these have a complex arrangement in total cellular complement of human satellite III. The analysis of human satellite III DNA on individual human chromosomes has been approached hy the use of rodent x human cell hybrids containing essentially only one whole human chromosome. Chromosomes examined to date include human chromosomes 1, 7, Il, 15 22 and the X. A combination of restriction enzyme digestion, gel electrophoretic and nucleic acid hybridisation techniques shows that the arrangements of the satellite III sequences located on each of these chromosomes are different and consist of sub-patterns of the pattern that can be identified in total human satellite IlL The results suggest that the satellite sequences on different human chromosomes have evolved independently of each other, and that there has been little exchange of satellite sequences between chromosomes since satellite III arose. On a more practical level, the restriction enzyme digestion patterns of human satellite DNAs may serve to "type"individual human chromosomes.

THEDISTRIBUTION OF SATELLITE DNAs IN CLOSELY-RELATED GENOMES OF INSECTS AND A NOTE ON THE INDUCTION OF POLYTENISATION OF SATELLITES IN CALLIPHORA GABRIEL DOVER Department of Genetics,

Anindication of the evolutionary behaviour of satellite DNAs may he obtained from the comparison of" common" satellites in closely related species, and an indication of the role these sequences may have in chromosome integrity or activity may be obtained from the behaviour of satellites during development. A study (Barnes, Webb and Dover, Chromosome,inpress) on seven sibling species of the melazsogaster species subgroup of Drosophile has shown that there are more than 10 satellite DNA components that are distributed such that each species is unique in the spectrum of satellite DNAs that it possesses but that nevertheless confine each species to a phylogenetic arrangement based on polytene inversion studies (Lemeunier and Ashburner, Proc. Roy. Soc. B, 193, 275, 1976). Digestion of isolated satellites with restriction enzymes and cross- hybridisation reveals differences in higher order organisation of some satellites that are shared between two or more species. The analysis of satellite DNAs in species and subspecies (races) of Clossiea (Amos and Dover) reveals a distribution of satellites such that there are no obvious homologies of satellites between species and races despite the abundance of satellite DNAS in the group. The distribution of satellite DNA is confounded also by the presence of supernumerary (B) chromosomes in two of the three species subgroups within the genus (Southern and Pell, Chromosome, 44, 319, 1973). The differences in satellite DNA between closely.related species, are discussed in relation to the possible nature of the failure of germ-line development in interspecific hybrids in Drssophila and to the contrasting evolutionary changes that have occurred in these two groups of insects. An analysis of the Calliphora (Dover and Ribbert) has revealed a contrasting behaviour in replication of satellite DNAs during normal polytenisation of DNA sequences in salivary glands and malpighian tubules (selective under-replication of satellites) and during induced polytenisation in nurse ccli nuclei (co-replication of all sequences). This is discussed in relation to the process of polytenisation in somatic and germ-line tissues.

THECOMPOSITION OF SUPPLEMENTARY DNA IN FLOWERING PLANTS R. K. i. NARAYAN Department of Agricultural , University College of Wales, Aberyscwyth

Thedivergence and evolution of species within many genera of plants is often accompanied by large-scale variation in chromosomal DNA. Investigations in diploid species of Let /zyrus, Allium and Lotion show that the distribution of quantitative DNA changes is not at random 416 GENETICAL SOCIETY OF GREAT BRITAIN within and among the chromosomes of the complement. There is evidence to show that the supplementary DNA contributing to the variation within genera is of uniform composition and organisation. In the genus Lathjvrus each additional picogram of DNA within euchromatin is accompanied by the addition of 1.2 picograms in heterochromatin. Each additional picogram of non-repetitive DNA is accompanied by an increment of four picograms of moderately repetitive DNA. Cross reassociation among repetitive and among non-repetitive DNA fractions from different species show substantial divergence in DNA composition. In general divergence in DNA composition is correlated with nuclear DNA amount. The degree of divergence is of the same magnitude in both repetitive and non-repetitive fractions.

2.CHROMOSOMES N MAUGNANCY

OCCURRENCE AND SIGNIFICANCE OF CHROMOSOME ABERRATIONS IN MALIGNANT CELLS GORAN LEVAN Deportment of Genetics, University of Got henburg, Sweden

Inthe last few years there has been an enormous increase in the amount of data on chromosome aberrations in human neoplastic disease. Our surveys of this material clearly indicate that the overall aberrations are not scattered randomly over the human karyotype but tend to cluster to specific chromosomes, viz. Nos. 1, 3, 5,7, 8, 9, 14, 17, 21 and 22. To further explore the significance of this clustering effect, we have turned to experimental materials. In sarcomas and leukaemias of the rat, for instance, the tendency to clustering can be detected and, moreover, what chromosomes are affected in individual cases appears to be dependent on the inducing agent. Thus, in the case of DMBA-induced rat malignan- cies, chromosome No. 2 has been frequently involved in aberrations. Acute treatment of normal rat cells with this chemical—in vivo or in vitro—has resulted in increased incidence of breakage and sister chromatid exchange preferentially in chromosome No. 2. We have looked upon these effects as indications of selective interaction between the carcinogen and genetic material in specific chromosomes. It may be speculated that in some cells this interaction will result, not in breakage but in sub-microscopic change causing malignant transformation. Once this change has taken place, secondary chromosome aberrations favouring rapid cell proliferation will accumulate, increasing the malignant capacity of the cells. These secondary aberrations mainly involve the chromosomes originally affected by the carcinogen and may consist of chromosomal events leading to increase in the relative amount of the chromosome segment underlying the transformation. Conversely, observation of the chromosomal changes may give a hint as to the sites in the karyotype of genie material important in the malignant development.

CLONESAND CHROMOSOMES IN HUMAN MALIGNANCIES SYLVIA D. LAWLER Deportment of Cyto genetics and Immunogenetics, Institute of Cancer Research and the Royal Morsden Hospital, London SW3 6JJ

Detectablechromosomal abnormalities are not a universal feature of human malignancy; the frequency exhibited varies from 50 per cent in acute myeloid leukaemia (AML) to over 90 per cent in the case of Philadelphia chromosome (Ph') positive chronic myeloid Ieukaemia. Karyotypic abnormalities of malignant cells may be clonal or non-clonal. For example, in acute lymphoblastic leukaemia the presence or absence of hyperdiploidy rather than the detection of dominant clones is the important feature. In AML non-random abnormalities peculiar to particular cytological types of disease are found in a minority of cases and, when therapy is successful, these abnormal clones are eliminated from the bone marrow. The progression of any malignancy may be accompanied by chromosomal evolution in a dominant clone or clones. Such changes may have diagnostic significance if detected before meta- morphosis occurs. Pre-malignant conditions such as Ph' positive leukaemia in its chronic GENETICAL SOCIETY OF GREAT BRITAIN 417 phase of monosomy 7 in the pre-leukaemic phase may be accompanied by additional changes when the leukaemia becomes acute. Attempts are being made to relate the phenotypic effects of the involvement of particular chromosomes in malignancies with the gene map. The question of unicellular or multicellular origin of tumours can also be studied bio- chemically. Up till now, the evidence has rested on studies of the sex-linked polymorphic enzyme systems. Immunological detection of cell surface marker characteristics has opened up a new approach to the study of cell lineage. Combination of cytogenetic, biochemical and immuno- logical approaches may lead to an elucidation of the importance of the clonal concept in human tumours.

THEANALYSIS OF MALIGNANCY BY CELL FUSION E. SIDEBOTFOM Sir William Dunn School of Pathology, Oxford

Alarge number of different types of hybrid cells have been made by fusing mouse tumour cells with diploid mouse cells. The tumour cells have included carcinomas, sarcomas, lymphomas and melanomas; some from spontaneous tumours and others experimentally induced with chemicals or virus. In all cases it has been found that the hybrid cells are either non-malignant or considerably less malignant than the tumour cell parent from which they are derived. It appears that the normal diploid genome is able to suppress malignancy. However, in many cases it has been found that the malignant phenotype reappears in hybrid cells after a variable period of culture in vivo or in vitro. The malignant segregants have always lost chromosomes. By using cells with appropriate cytogenetic or enzyme markers it has been shown that in some of the systems tested all malignant segregants had eliminated both copies of chromosome 4 derived from the diploid mouse cell. The elimination of both copies of this chromosome was, however, not in itself sufficient to ensure the reap- pearance of the malignant phenotype; some hybrids without a diploid chromosome 4 did not generate progressive tumours. An additional event that could not be related to elimina- tiori of a specific chromosome is evidently required. Biochemical studies of the membrane glycoproteins of non-malignant hybrid cells and malignant segregants derived from them have revealed a consistent difference in the poly- saccharide moiety of one specific glycoprotein. This difference has been detected in a wide range of different tumours and it remains linked to the malignant phenotype in a test which allows non-malignant cells to be selected from tumour cell populations by the use of the lectin wheat germ agglutinin. (Most of the work reported above was carried out by H. Harris, M. E. Bramwell and J. Jonasson.)

ACYTOGENETIC STUDY OF THE INTERACTIONS OF ADENOVIRUS AND RAT CELLS, INFECTED AND TRANSFORMED P. H. GALLIMORE Department of Cancer Studies, University of Birmingham Medical School, Birmingham BIS 2T3

Asa consequence of permissive infection, adenovirus 12 (Ad-12) produces specific cytogenetic damage to human autosomes I and 17. My presentation will be concerned with the cytogenetic changes observed in rat embryo cells infected with oncogenic Ad-12 (non-permissive for rat cells) and non.oncogenic Ad-2 (semi-permissive for rat cells). Both of these human adenovirus serotypes morphologically transforms rodent cells in tissue culture. The cytogenetic status of a number of independently derived Ad-2 and Ad- 12 transformed rat embryo cell lines has been obtained using a Giernsa banding technique. Thus far, all of the Ad-2 transformed lines analysed have marker chromosomes and show stem line proliferation. The origins of some of these marker chromosomes will be defined and the relevance of cytogenetic abnormalities to tumorigeni- city in this experimental system is discussed. 418 GENETICAL SOCIETY OF GREAT BRITAIN 3.GENERAL CONTRIBUTED PAPERS

THE MEANING OF GENE CONVERSION SPECTRA B. C. LAMB and A. GHIKAS Botany Department, Imperial College, London SW7 2BB

Leblon(Molec. gen. Genet., 115, 36, 1972 and 116, 322, 1972) defined four kinds of con- version spectrum: type A, rare postmeiotic segretation (PMS), excess of conversion to wild- type; type B, rare PMS, excess of conversion to mutant; type C, PMS not rare, excess of conversion to wild-type; type D, PMS not rare, excess of conversion to mutant. In Ascobolus immersus he found a clear relation between a mutation's conversion spectrum type, its molecular type (addition or deletion frame-shifts, or base substitutions) and the mutagen used. In the Pasadena strains of Ascobolus immersus we have studied the conversion properties of nine UV, nine NG (nitrosoguanidine), eleven ICR 170 induced and nine spontaneous white ascospore mutations. Each mutant was crossed to three different derived wild-type stocks. These crosses gave an almost continuous spread, from 0-100 per cent, for PMS frequencies and for relative frequencies of conversion to wild-type and conversion to mutant, so spectrum types A and B intergraded, as did C and D, and A and B intergraded with C and D. Spontaneous and UV-induced mutations gave types A, A/B, C, D and C/D, while ICR 170 gave A, B, A/B, C, D and C/D ,with B and C most frequent. Individual mutations often gave different conversion spectrum types when crossed to the different derived wild- types, e.g. A in one cross, D in another. The meaning of conversion spectra therefore needs re-evaluating. Because of asymmetric hybrid DNA formation, a distinction must be made between conversion spectra and correction spectra. One should also consider excision- repair triggered by mispairs from heterozygous cryptic mutations (Lamb, Molec. gen. Genet., 137, 305, 1975) near to the conversion site, as well as repair triggered at the site itself.

THEINHERITANCE OF BARBITURATE SLEEPING TIME IN MiCE D. P. LOVELL Medical Research Council, Laboratory Animals Centre, Woodmansterne Rood, Carshalton, Surrey SMS 4EF

Thelength of anaesthesia following hexobaritone injection into mice is affected by both genetic and environmental factors (E.S. Vessel, Pharmacology, 1, 7, 1968). A 6 x 6 diallel cross where pentobarbitone-induced sleeping time was measured has been carried out. Animals were fed either a commercial mouse diet or a protein-deficient diet, and tested at one of three different temperatures. The environmental factors were imposed on the diallel in a factorial analysis of variance. A considerable number of genotype x environmental interactions occurred and the form of inheritance of the character was predominantly additive gene action with no evidence of epistasis.

GENETICCONTROL OF QUANTITATIVE ANTIBODY RESPONSE TO BACTERIOPHAGE MS2 IN MICE N.NGWA SUH,A. EBPJNGERand B.BAINBRIDGE Immunology Unit, Departments of Biochemistry and Microbiology, Queen Elizabeth College, London

Thegenetic control of the secondary immune response to the antigen MS2 bacteriophage, in presence and absence of adjuvant, was investigated in 12 different strains of inbred mice, consisting of 227 test and 94 control animals. Bacteriophage MS2 (titre 1 x l0' pfu/ml), diluted, 1 : 10 in saline, was injected either intraperitoneally in 0.1 ml doses into female mice or emulsified in incomplete Freund's adjuvant and then injected interperitoneally into male mice. Both male and female mice were boosted on day 21, such that both sets of animals received the same total dose of MS2 antigen and all were bled out 10 days later. Using a '251-labelled MS2 antigen excess assay, high antibody responses were observed in adjuvant-injected mice, compared to mice injected with antigen alone. In the adjuvant GENETICAL SOCIETY OF GREAT BRITAIN 419 group, high antibody responses were obtained in strains Sc.Sn, BIOA, BIOBR and BALB/C, and low responses in strains ASW, DBA/2 and SWR/J. In the saline-injected group, strains ASW, DBA/2, SWR/J and BALB/C gave relatively high responses compared to strains Sc.Sn, B1OA and B1OBR, thus showing opposite behaviour compared to adjuvant-treated animals, except for BALB/C mice. Although the immune response appears to be related to the genetic composition of each strain, high and low responder status depends also to some extent on mode of immunisation but the mechanism mediating such response is at present unknown.

SELF-TOLERANCEAND THE Fl HYBRID RESPONSE R. SUBRAMANIAN and A. EBRINGER Immunology Unit, Department of Biochemistry, Queen Elizabeth College,London Ithas been suggested that self-tolerance is acquired during the early life of a vertebrate animal by elimination of self-reactive clones. This theory has been tested by reviewing the Fl hybrid response in one-way mixed leucocyte culture (MUL) systems. The response was compared using Fisher's dominance index (D), Fl—L D= —l (H—L) where H, Fl and L are the allogeneic, Fl hybrid and syngeneic parental thymidine uptakes respectively. Since both Fl hybrid and syngeneic responder cells share identical antigens with the stimulator cells, the Fl hybrid response would be expected to approach that of the syngeneic background response and give a value of 1 for the dominance index. In a review of 79 one-way MLC systems, consisting of 65 human and 14 murine combina- tions, the mean dominance index was found to be+0.230±0.llO (mean±S.E.), a value consistent with co-dominant inheritance (p <0.001) and similar results have been obtained with immunogenetic (IR-gene) systems. This result suggests that self-tolerance does not occur by a gene product or protein recognition event during the early life of an animal, but by an active process of gene or nucleic acid recognition during an immune response.

STUDIESON MEIOTIC DRIVE IN THE MOSQUITO, AEDES AEGYPTI (L.) M. E. NEWTON, R. J. WOOD and D. I. SOUTHERN Department of Zoology, University of Manchester InAedes aegypti, sex-ratio distortion in favour of males is determined by the male parent and is the outcome of meiotic drive involving the Y-borne M (sex) and D (distorter) loci of linkage group I. In the presence of D, X chromosomes which are sensitive to distortion undergo preferential fragmentation during male with a consequent reduction in the proportion of functional X-bearing spermatozoa. The effect is seen in an extended range of DNA levels within spermatozoa of distorting males. These include values up to 4C levels as well as a few with less than the IC amount. The separate identities of M and D have been established and their positions in chromosome I have been determined in relation to Giemsa C-bands.

PHOSPHOGLUCOMUTASE PHENOTYPES DETECTED BY ISOELECTRIC FOCUSING: POPULATION FREQUENCIES, FAMILY STUDIES AND LINKAGE ANALYSIS C.M.WEST, N. D. CARTER, E. EMES and B. H. PARKIN Department of Child Health, St. George's Hospital Medical School, London, and The Metropolitan Police Forensic Science Laboratory, London

Phosphoglucomutase(PGM) polymorphism in man has been extensively studied in starch gel electrophoresis, and interest in the enzyme has recently been revived by separation 420 GENETICAL SOCIETY OF GREAT BRITAIN of phenotypes in isoelectric focusing. Using this new method it is possible to distinguish four common alleles at the PGM, locus (PGM 1+, PGM I —,PGM2+, PGM 2—) which code for 10 phenotypes, previously recognized as only three electrophoretic forms, coded by two alleles. Family studies show that with a few exceptions (to be described) these four alleles are inherited in a Mendelian fashion. We tentatively propose a mechanism involving two mutations and a cross-over for the evolutionary origin of the four alleles. Studies on several chimpanzee lysates showed a PGM, pattern identical to that produced by the PGM l+l+genotype in man. This latter form is the one predominant in all human populations studied to date, and this PGM 1 + may be the ancestral PGM, allele in man. This new system provides useful information for linkage and population studies. A large extended family of 1500 people in Newfoundland is being studied for genetic association with a high incidence of lymphoma and alleles at the gene loci for Rhesus and PGM, have been analysed for linkage using the technique of isoelectric focusing to determine PGM1 phenotypes. The new data confirm previous reports that weak linkage is present. PGM, gene frequencies in the Newfoundland community were: 1—, 0l53; 1+, 0537; 2—, 0076; 2+, 0233. These may be compared with a Northern European community from London (total 2773) which gave 1—, 0l32; 1+, 0627; 2—, 0.058; 2+,0l82. The gene frequencies in a Negro population residing in London (384) were I —,0l26;1+, 067l; 2—, 0029; 2+, 0l73: and in an Asian group in London (120 studied): 1—, 0096; 1+, 0537; 2—, 0l2l, 2+, 0246. The significance of these polymorphic frequences and of some rare PGM phenotypes will be discussed.

AUTOSOMALDOMINANTLY INHERITED ADDUCTOR LARYNGEAL PARALYSIS—A NEW SYNDROME WITH A POSSIBLE LINKAGE TOHLA MARGARET MACE', ELSPETH WILLIAMSON' and D. WORGAN' 1 Department of Biology, University of Southampton, 'Department of Child Health, Southampton General Hospital, Southampton, and 'E.N.T. Department, Royal South Hants Hospital, Southampton

Afamily with hereditary, congenital, bilateral adductor paralysis of the larynx and no other clinical abnormality is reported. Four males and one female with this disorder, covering three generations of this single family, were seen and the condition appears to be transmitted as an autosomal dominant. This disorder has not been described previously. A search for linkage in this family with the loci for 19 other genetic marker systems were conducted. Very close linkage with several genetic markers could be excluded. There was a possibility of linkage with HLA and GLO. With HLA for phase known offspring there were three non-recombinants and no recombinants and the peak male lod score (+ 1.505) at zero recombination gave a probability of 32 : I in favour of linkage. We suggest the locus for this new disorder may be on chromosome 6, closely linked to HLA and GLO.

CORRELATIONBETWEEN CHROMOSOMAL ABERRATIONS AND SISTER CHROMATID EXCHANGE B.BHATT Department of Genetics,

Neitherthe mechanism of the formation of chromosomal aberrations, nor the mechanism " of sister chromatid exchange is known. Aberrations seem to result from "mis-repair of damaged DNA while sister chromatid exchanges (SCE) are a replication-dependent pheno- menon. Mutagens that induce aberrations during or before S-phase of the cell cycle cause a dramatic increase in SCE. A parallel distribution of chromatid breaks and SCE induced by DMBA and TMBA has been reported by Ueda etal.(Xature, 262, 581-583, 1976), but this correlation is disputed by Ikushima (Nature,268,235-237, 1977). Assuming that SCE are a replication-dependent phenomenon one could study the possible correlation between SCE and chromatid breaks induced prior to replication. Such a corn- GENETICAL SOCIETY OF GREAT BRITAIN 421 parison between breaks and SCE should provide useful insight into the mechanism of SCE. CHO-Kl (chinese hamster ovary) cells were labelled with 5-bromodeoxyuridine (BUdR) according to the FPG technique (Perry and Wolff, Xature, 251,156-158,1974). The cells were treated with mitomycin C at the beginning of the second cycle. The resulting chromatid breaks and SCE were compared either directly or on photographs. The implications of these results will be discussed.

QUANTITATIVEANALYSIS OF THE FREQUENCY OF SATELLITE ASSOCIATION IN HUMAN ACROCENTRIC CHROMOSOMES AFTER EXPOSURE TO LOW DOSES OF GAMMA RAYS M. T. KHOKHAR, T. MN and J. A. HOUGHTON Cytogenetics Unit, Department of Microbiology, University College, Gaiway, Ireland

Ithas been suggested that satellite association of acrocentric chromosomes may be one of the determining factors in the causation of non-disjunction in man. Down's syndrome, the most common human autosome disorder, is usually caused by non-disjunction of chromo- some 21. Prospective and retrospective studies have shown that the incidence of Down's syndrome may be higher among the children of parents who have been exposed to ionising radiation. To determine whether irradiation leads to an increased incidence of satellite association and, therefore, an increased risk of chromosome non-disjunction, the frequency and patterns of satellite associations in metaphase preparations from irradiated blood samples were investigated. Samples were exposed to small doses of 60Co gamma rays and associated complexes were evaluated using the criteria of Zang and Back (Gytogenetics, 7, 455, 1968). Blood samples from normal and Down's syndrome individuals, parents of Down's and parents of advanced age of normal children were irradiated at room temperature and at 37°C. Cultures were harvested after 48 hours at 37°C and 1000 metaphases at each dose level were analysed. The frequency of cells showing either 0-4 association complexes were compared between irradiated and unirradiated, control samples. There is no apparent indication of a dose-response relationship for an effect of irradiation on the frequency of satellite association within the dose range used. Similar results were obtained when satellite association was determined using the silver staining technique which permits the identification of physical connectives between the associated acrocentric chromosomes. These results and their relevance to chromosome abnormality in man will be discussed.

ARYLHYDROCARBON HYDROXYLASE INDUCIBILITY IN LUNG CANCER A. E. H. EMERY, R. ANAND and N. DANFORD Department of Human Genetics, University of Edinburgh

Lungcancer is largely caused by smoking, but since all smokers do not develop cancer it may be that certain individuals are constitutionally predisposed to the disease should they smoke. Polycyclic aromatic hydrocarbons (PAN) in tobacco smoke are broken down by the enzyme aryl hydrocarbon hydroxylase (AHH) to carcinogenic epoxides. AHH activity is induced by a variety of substances including PAN. We have studied AHH inducibility in peripheral blood lymphocytes in patients with established squamous-cell lung cancer, patients with cancers at sites other than lung and controls matched for age, sex, social class and smoking habits. All enzyme assays were carried out "blind "withoutknowledge as to the source of the material. The proportion of high inducers was significantly greater among patients with lung cancer (but not patients with other cancers) than among controls. This confirms the original suggestion by Kellerman et at. (New Engi. J. Med., 289, 934, 1973) that, besides smoking, a constitutional factor may also be involved in the pathogenesis of lung cancer. However, our studies of AHH inducibility among relatives of lung cancer patients suggest that this is not inherited in any simple manner and may have a multifactorial basis. 422 GENETICAL SOCIETY OF GREAT BRITAIN

MEIOTICAND RADIATION STUDIES IN FOUR OLIGOCHIASMATIC HUMAN MALES

ANN C. CHANDLEY, E. THOMSON and J. FLETCHER MRC Clinical and Population CytogeneticUnit,Western General Hospital, Edinburgh, EH4 2XU

Ina few cases of human male sterility, aberrant meiotic development, possibly attributable to the action of a mutant gene, is observed. Although such individuals are phenotypically normal and have a normal somatic karyotype, their chromosomes display synaptic irregu- larities at meiotic prophase and, at diakinesis, low chiasma counts and high numbers of univalents are recorded. In extreme cases, spermatocytes may be almost totally achiasmate and complete germ cell maturation arrest and azoospermia are found. Radiation studies carried out on peripheral blood lymphocytes from one such azoospermic individual having a mean chiasma count of nine per cell at diakinesis, indicated a reduced facility for repairing chromosomal DNA (Pearson et al. Cylogenetics, 9, 460, 1970). It was suggested that the repair defect could also explain the failure of chiasma formation at meiosis. However, recently completed studies on four oligo-chiasmatic males, carried out in our own laboratory, give no such indication of reduced repair capacity following irradiation to blood lymphocytes or fibroblasts. It would appear that the "low chiasma count "conditionin sterile men may be of mixed aetiology.

LATEDNA REPLICATING PATTERNS IN THE CHROMOSOME COMPLEMENT OF MAN AND THE GREAT APES (PAN TROGLODYTES, GORILLA GORILLA AND PONGO PYGMAEUS) HECTOR SEUANEZ MRC Clinical and Population Cytogenetics Unit, Edinburgh

LateDNA synthesis in human and great ape chromosomes was studied using the thymidine analogue 5-bromodeoxyuridine (BUdR) following two different protocols.(i) Peripheral blood lymphocytes were grown in culture medium containing BUdR for the first 43 hours, and then in medium containing thymidine for the final 5 hours before harvesting. (ii) Peripheral blood lymphocytes were grown in medium containing thymidine for the first 68 hours, and then in medium containing thymidine for the final 5 hours before harvesting. Late DNA replicating regions in the chromosome complement of man and the great apes coincided with regions of positive Q-,G-and C-banding in (i), and with R-band regions in (ii). A comparison between the late replicating pattern of chromosomes recognised as "inter- specific homologues" (Paris Conference 1971; supplement 1975) showed remarkable similarities between human and great ape chromosomes. The evolutionary conservation of the late DNA replicating pattern in the chromosomes complement of man and great apes will be discussed, and compared to similar studies in other phylogenetically related species.

AUTORADIOGRAPHICANALYSIS OF DNA REPLICON SYNTHESIS IN THREE CANCER-PRONE CONDITIONS C. H. OCKEY and A. M. R. TAYLOR Paterson Laboratories, Christie Hospital, Manchester, and Deportment of Cancer Studies, University of Birmingham

Severalof the inherited cancer-prone syndromes in man have been shown to be sensitive to various mutagenic agents and to possess defects in their DNA repair mechanisms: ataxia telangiectasia (y-irradiation), xeroderma pigmentosum (UV irradiation), Fanconi's anaemia (mitornycin C). In Bloom's syndrome on the other hand defects have been observed in the DNA synthesis mechanism of cultured cells (Hand and German, Hum. Genet., .98, 297, 1977; Giannelli et al., .ftfature, 265, 466, 1977). Using a quantitative system of analysis, replication synthesis from DNA autoradiographs GENETICAL SOCIETY OF GREAT BRITAIN 423 of pulse labelled cells was investigated in fibroblast cultures of dermal cells from ataxia telangiectasia, basal cell naevus and Bloom's syndrome. In all cases the parameters—rate of chain elongation, distance between initiation sites and the compensatory relationship between those two factors—were similar. There was no suggestion of any defects in DNA synthesis when cells were grown at 2-4 x 1 0°/cm5. Measurements of cell cycle parameters in cultures of Bloom's syndrome fibroblasts indicated that cell density was an important factor in altering DNA synthesis parameters. At low cell densities 2-6 x l03/cm2 the S period was extended compared to control cells, while at densities of 2-3 x 10°/cm2 the S period was of normal duration. It is suggested that this factor contributes to the low rate of DNA chain growth observed in Bloom's syndrome.

INCREASEIN CHROMOSOMAL RADIOSENSITIVITY OF CHINESE HAMSTER CELLS WITH IN VITRO PASSAGE CAROL Y. LYONS, D. SCOTT and JANE R. CONNELL PatersonLaboratories,Christie Has pita! and Holt Radium Institute, Manchester M20 98X

Cellsfrom cancer-prone individuals with a trisomic chromosome constitution (e.g. Down's syndrome) have an elevated sensitivity to X-ray-induced chromosome aberrations (Sasaki et al., Mutation Res., 10, 617, 1970). The sensitivity of a trisomic Chinese hamster cell line (passage 40) to X-ray-induced chromosome damage was found to be twice that of the diploici line (passage 16) from which it arose spontaneously. A line carrying a simple inversion (passage 9), was of similar sensitivity to the diploid. Further experiments suggest, however, that the difference in radiosensitivity is not related to the karyotypic differences but to age in culture since the diploid cells were found to increase in sensitivity between passage 16 and 47. Cells may lose their repair capacities with increasing in vitro culturing.

4.CLINICAL GENETICS SOCIETY

DIAGNOSIS OF LYSOSOMAL STORAGE DISEASES BY LEUCOCYTE ENZYME ASSAY A. COOPER, B. FOWLER and I. B. SARDHARWALLA Willink Biochemical Genetics Unit, Royal Manchester Children's Hospital, Pendlebury, Manchester

Wereport here our approach to investigation of lysosomal storage diseases in suspected patients. These disorders are usually characterised by a specific enzyme deficiency and often exhibit classical features. However, for many of these diseases, variant or atypical cases exist which do not fit the classica' picture. In this Unit we assay 15 enzymes on a single blood sample for all referred patients. The following enzymes are assayed in a Triton Xl00 extract of leucocytes obtained from 10 ml of blood; Total -Hexosaminidase (Sandhoff disease); fl-Hexosaminidase A (Tay- Sachs disease); -Glucuronidase (Sly's disease); -Galactosidase (GMI Gangliosidosis); Aryl Suiphatase A (Metachromatic leucodystrophy); -Mannosidase (Mannosidosis); ce-Fuco- sidase (Fucosidosis); a-Galactosidase (Fabry's disease); ce-Glucosidase (Pompe's disease); -G1ucosidase (Gaucher's disease); 8-Galactoscerebrosidase (Krabbe's disease); Sphingo- myelinase (Niemann-Pick disease A and B). Acid phosphatase (Total Acid Phosphatase deficiency), ss-Mannosidase (Mucolipidosis II and III) and o-N-acetyl-glucosaminidase (Sanfihippo B) are assayed in plasma. Optimal conditions have been established for rapid microassays using commercially available fluorogenic or chromogenic substrates. High sensitivity has been obtained by using four methylumbelliferyl conjugates where applicable. To date we have investigated more than 700 patients of whom 16 were affected by a specific lysosomal enzyme deficiency. In seven of these affected patients the specific enzyme defect had been clearly predicted by the clinical features. However, in the other nine the exact enzyme defect had not been suspected on clinical grounds. The approach of assay of the majority of the enzymes deficient in lysosmal storage diseases allows detection of clinical variants. In case of neurological degenerative disorders of unknown aetiology a finding of normal enzyme levels allows exclusion of many of the lipid storage diseases. 424 GENETICAL SOCIETY OF GREAT BRITAIN

MAYSPINA BIFIDA RESULT FROM AN X-LINKED DEFECT IN A SELECTIVE ABORTION MECHANISM? J. BURN and D. GIBBONS Department of Medicine, Royal Victoria Inflrmary, Queen Victoria Road, Newcastle upon Tyne Itis proposed that the major genetic factor in determining the birth of children with neural tube defects is a single X-linked gene. It acts as an X-linked dominant, not by producing neural tube defects, but by enabling the affected foetus to survive selective spontaneous abortion. This mechanism, mediated at the deciduo-placental junction, could be under the control of both maternal and foetal genes. With more defective alleles, survival would become more likely, reaching a maximum in the homozygous affected female, foetus of a homozygous affected mother. The female excess in anencephaly is greater than that in spina bifida, due to its prenatal severity, thus requiring relatively more mutant alleles for survival. This hypothesis is an extension of the multifactorial model and will be shown to be compatible with previous research data on the aetiology of spina bifida, including the recent work on prior abortions. The work of one of us (D. G.) will be used to briefly illustrate a possible biochemical pathway within the placenta.

PARENTALCONSANGUINITY AND FOETAL GROWTH J. R. SIBERT, M. JADHAV and S. IMBARAJ University Hospital of Wales, Cardiff, and Christian Medical College Hospital, Vellore, South India

Anthropometricmeasurements made on 322 newborn infants in South India were related to parental consanguinity. Uncle-niece and first-cousin marriages were common and the average coefficient of inbreeding as high as 00329. The measurements (weight, length, head circumference and triceps and subscapular skinfold thickness) of the uncle-niece group (52 infants) were smaller than those of the first-cousin group (61 infants) which in turn were smaller than the non-consanguinous group (196 infants). Statistical significance (P <0.01) was only recorded, however, between the weights of the three groups (means 26504 g, 2794'l g, 2853'O g) and between the lengths of the uncle-niece group and the non-consanguinous group (means 46'92 cm and 4779 cm). There were no social class or residential differences between the groups. We conclude that there are likely to be recessive genes present in the population slightly retarding foetal growth.

ANUNUSUAL COINCIDENCE M. SEABRIGHT and N. GREGSON Wessex Regional Cytogenetics Unit, General Hospital, Salisbury Thisis a report of an unusual coincidence following amniocentesis and termination of the pregnancy X-linked muscular dystrophy associated with testicular feminisation.

HLAAND 21 HYDROXYLASE DEFICIENCY R. HARRIS, P. T. KLOUDA and D. A. PRICE Department of Medical Genetics, St. Mary's Hospital, Manchester, and Royal Manchester Children's Hospital, Pendlebury, Manchester Thelocus for 21 -hydroxylase deficiency has been shown to be closely linked to the HLA-B locus of the major histocompatibility complex (MHC) by studying 25 families with one or more children with 21-hydroxylase deficiency. Both salt losing and non-salt losing 2 1-hydroxylase deficiency appear to be linked to HLA. HLA typing may become of great practical value within families in the early identification of potentially affected individuals and in distinghishing heterozygotes from homozygotes. GENETICAL SOCIETY OF GREAT BRITAIN 425

ASURVEY OF CONGENITALLY DEAF CHILDREN IN THE NORTH-WEST OF ENGLAND IAN G. TAYLOR Department of Audiology and Education of the Deaf, University of Manchester

Theimportance of identifying accurately the aetiological factors in congenital sensori- neural deafness is self-evident. Large studies of congenital sensori-neural deafness indicate that one-third of these have no identifiable aetiological factor. The most important large study, by Fraser, indicated that a considerable proportion of the so-called "unknown" group were likely to have a genetic basis. The study to be reported concerned the detailed examination of 86 children in one school " for the deaf. The "familialgroupwere compared with the" unknown" group in respect of hearing loss and incidence of sensori-neural deafness in the families. Two approaches were made to the analysis of the data, one audiological and the other through Mendelian-segregation analysis, the hypothesis to be considered being that the whole of this group of children of unknown aetiology are in fact isolated cases of transmission of deafness through an autosomal recessive gene.

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