Effects of exposing Drosophila melanogaster parents to ethanol on expression of vestigial in their progeny

When parental Drosophila melanogaster were chronically exposed at 28 degrees C or 24 degrees C to ethanol during their larval and pupal stages of development, their progeny, produced when parents were 5-16-day-old adults, showed modified expression of vestigial alleles in heterozygous and homozygous combinations. Parental alcohol effects were dependent on parental rearing temperature. We conclude that parental environment (alcohol, temperature) causes heritable but transitory changes in progeny phenotype that are elicited by exposure of germ cells to alcohol.     

Methods to predict transgressive segregation in barley and other self-pollinated crops

Most of agronomically important characters are biometric traits. An improvement of these traits in cultivated plants by deriving segregants superior to parents, which could be developed as cultivars, is a main goal in breeding of self-pollinated crops. Two problems need to be solved: when will the progeny be better than its parents and how can a genetic potential of a given pair of parental genotypes be predicted? In this paper, transgressive segregation in homozygous barley populations is shortly reviewed. Various approaches to choosing parental forms are shown, and a theoretical method for predicting the frequency of transgressive segregants in a homozygous population is presented. Additionally, relationships between parental diversity estimated with molecular markers and the progeny performance are discussed. Although the prediction of transgressive segregation is still a problem, it seems promising to apply an approach measuring the performance of the parental genotypes and estimating their genetic distance by molecular markers.     

Mating behavior of Phytophthora parasitica: Evidence for sexual recombination in oospores using DNA restriction fragment length polymorphisms as genetic markers

Cloned nuclear DNA fragments that detected restriction fragment length polymorphisms (RFLPs) in homozygous loci of isolates of Phytophthora parasitica were used as genetic markers to investigate sexual recombination during oospore formation. It was found that the majority of the 23 oospore progeny studied in each of the two crosses carried both of the parental markers. However, aberrant recombination patterns were observed; some of the progeny were homozygous at one RFLP locus, whereas at another locus both of the parental markers were present. Only two of the progeny of each cross did not show sexual recombination with any of the four or five RFLP markers used. Mitochondrial DNA (mtDNA) was uniparentally inherited. In both crosses the majority of the progeny carried the mtDNA type of one of the common parental strains.     

In vitro assay to select rainbow trout with variable resistance/susceptibility to viral haemorrhagic septicaemia virus

The merit of a candidate criterion of resistance to viral haemorrhagic septicaemia virus (VHSV) was tested with the view of producing experimental trout progeny with a predictable level of resistance. The criterion, the measure of in vitro viral replication in excised fin tissue (VREFT) was previously developed. Three experiments were performed, using both ordinary and homozygous doubled-haploid breeders. A set of 48 progeny was tested. Breeders were individually scored for repeated measures of VREFT, and the progeny were tested against VHSV (strain 07-71, serotype 1) through a waterborne challenge (5 x 10(4) pfu ml(-1) during 2 h). Analysis of repeated measures of VREFT revealed the risk of identifying 'false' resistant individuals. The highest value should be considered the most predictive of the resistance status. Survival of progeny ranged from 0 to 100% according to the group and the experiment. The survival was correlated to the mean VREFT value of the breeders in Expts 1 and 2 (R = 0.96 and 0.61 respectively), but not in Expt 3 (R = 0.36, ns) where all tested progeny were highly susceptible. Results thus indicate that viral growth in fin tissue is genetically correlated to resistance to waterborne disease and may be used to produce selected progeny, at least at the experimental scale. Possible implications of the relationship between VREFT and resistance for the study of resistance mechanisms are discussed.     

Long homozygous chromosomal segments in reference families from the centre d'Etude du polymorphisme humain

Using genotypes from nearly 8,000 short tandem-repeat polymorphisms typed in eight of the reference families from the Centre d'Etude du Polymorphisme Humain (CEPH), we identified numerous long chromosomal segments of marker homozygosity in many CEPH individuals. These segments are likely to represent autozygosity, the result of the mating of related individuals. Confidence that the complete segment is homozygous is gained only with markers of high density. The longest segment in the eight families spanned 77 cM and included 118 homozygous markers. All individuals in family 884 showed at least one segment of homozygosity: the father and mother were homozygous in 8 and 10 segments with an average length of 13 and 16 cM, respectively, and covering a total of 105 and 160 cM, respectively. The progeny in family 884 were homozygous over 5-16 segments with average length 11 cM. The progeny in family 102 were homozygous over 4-12 segments with average length 19 cM. Of the 100 individuals in the other six families, 1 had especially long homozygous segments, and 19 had short but significant homozygous segments. Our results indicate that long homozygous segments are common in humans and that these segments could have a substantial impact on gene mapping and health.     

A targeted screen to detect recessive mutations that have quantitative effects

A key problem in mutagenesis research is developing methods that are sufficiently sensitive to detect a wide range of abnormal phenotypes. Major variants may be easy to identify, but it can be difficult to detect mutations that have subtle effects, particularly on a complex genetic background. This paper describes a targeted mutagenesis protocol with enough sensitivity to detect recessive mutations that have modest quantitative effects. The procedure relies on consomic inbred strains of mice—strains in which one homologous pair of chromosomes of an inbred strain has been replaced with the corresponding pair from a donor strain. Mice that carry the desired donor chromosome—the target of the screen—are mutagenized and bred back to the original recipient strain. The first-generation progeny (G1) that are heterozygous only for the donor chromosome are also bred back to the recipient strain. G2 animals that inherit nonrecombinant donor chromosomes are identified by genotyping. These animals may be backcrossed repeatedly to the recipient strain to dilute off-target mutations, but ultimately, nonrecombinant G2 animals are bred to each other. Their G3 progeny are genotyped at markers spaced at 5- to 10-cM intervals to identify mating pairs that are homozygous for shared segments of the mutagenized donor chromosome. Entire litters of G4 progeny that are homozygous for defined intervals are screened. By comparing phenotypes within and among litters of nearly isogenic G4 animals, mutations can be verified and simultaneously mapped with a precision of 5–10 cM. This method has the potential to consistently detect mutations that have effects on trait means of well under one standard deviation. Received: 29 December 1998 / Accepted: 17 March 1999     

On the Genetic Control of Genes Located in the Sex-Chromosome Heterochromatin of DROSOPHILA MELANOGASTER

The genetic properties of a recessive autosomal point mutant, "daughterless" (symbol: da), are described. They are: (1) da maps in the euchromatin of 2L at about position 39 on the genetic map and between 27D and 31E on the salivary map; (2) when homozygous in females, da causes the production of unisexual male progenies owing to sex differential zygote mortality in the egg stage; (3) da has no effect on the progeny of mutant males; (4) female zygotes die, while at least some male zygotes survive, irrespective of the number of Y chromosomes or the amount of X-chromosome heterochromatin carried by either the mutant female or her progeny; (5) homozygous da progeny of heterozygous parents also show sex differential survival favoring males indicating a da effect in development as well as in oogenesis, but with the developmental effect much less pronounced; (6) extra heterochromatin from either the X or Y chromosome in either the mother or her progeny can reduce the mortality caused by da in development (and, therefore, conceivably can also reduce the abnormality in oogenesis, although this is obscured by the severity of the maternal effect.)—From these results, it is suggested that (1) da regulates either the activity of structural genes in the sex chromosome heterochromatin or the activity or stability of their products, and (2) it is a defective product of sex chromosome heterochromatic genes that causes the abnormalities resulting in the observed mortality of heterozygous progeny of homozygous mutant mothers and of homozygous mutant progeny of heterozygous mothers.—The striking parallels with the properties of the gene, "abnormal oocyte" are noted as is the close linkage between the two loci. The possibility of a special sex-chromosome-heterochromatin-regulator region on chromosome 2 is considered.     

Suppressor genes with gender differences in activity in natural populations of Drosophila robusta: another approach to wild-type

Homozygous or hemizygous expression of an X-linked wing mutant of Drosophila robusta varies from a rudimentary wing that does not reach the tip of the abdomen (called 'club') to forms with full-sized but curled or crumpled wings (called 'curly'). Homozygous club females crossed to flies from natural populations or laboratory stocks derived from wild flies invariably produce significantly less club male progeny than the 100% expected, most of them exhibiting less severe phenotypes: 'curly' forms and wild-type. The male progeny from similar crosses using curly females tend to be predominantly normal. By contrast, the male progeny of outcrossed females homozygous for an X-linked eye colour mutant, vermilion, are all vermilion. The data indicate that natural populations of D. robusta contain suppressors of the wing mutant but not of the eye colour mutant studied. Activity of the suppressors differs by gender: in experiments in which genetic theory expects similar results in the two sexes, males consistently show stronger effects of the suppressors than females.     

EXT gene family member rib-2 is essential for embryonic development and heparan sulfate biosynthesis in Caenorhabditis elegans

EXT gene family members including EXT1, EXT2, and EXTL2 are glycosyltransferases required for heparan sulfate biosynthesis. To examine the biological functions of rib-2, a member of the Caenorhabditis elegans EXT gene family, we generated a mutant worm lacking the rib-2 gene using the UV-TMP method followed by sib-selection. Inactivation of rib-2 alleles induced developmental abnormalities in F2 and F3 homozygous worms, while F1 heterozygotes showed a normal morphology. The F2 homozygous progeny generated from the F1 heterozygous hermaphrodites somehow developed to adult stage but exhibited abnormal characteristics such as developmental delay and egg-laying defects. The F3 homozygous progeny from the F2 homozygous hermaphrodites showed early developmental defects and most of the F3 worms stopped developing during the gastrulation stage. Whole-mount staining analysis for heparan sulfate using Toluidine blue (pH 2.5) revealed a defect of heparan sulfate biosynthesis in the F2 homozygotes. The analysis using fluorometric post-column high-performance liquid chromatography also uncovered reduced production of heparan sulfate in the rib-2 mutant. These results indicate that rib-2 is essential for embryonic development and heparan sulfate biosynthesis in C. elegans.     

Mutagenesis of a race-specific rust resistance gene in Antirrhinum majus using a transposon-tagging protocol

In a transposon tagging experiment, lines of Antirrhinum majus exhibiting both race-specific resistance (homozygous for the dominant R gene) to the rust pathogen Puccinia antirrhini and a high frequency of transposition were crossed with homozygous susceptible lines. From a screen of 11153 F₁ progeny, 15 individuals were detected showing susceptibility to rust race α. Six of these exhibited a susceptibility phenotype (classified as type B) not previously observed. A control experiment involving the same tagging strategy but employing lines that do not exhibit high rates of transposition did not yield any susceptible mutants from a screen of 6243 progeny. In experiments on the heritability and stability of the mutation, the six plants exhibiting susceptibility phenotype B produced progeny in which the R locus had reverted to an active form (i.e. some of the progeny were resistant), a classic characteristic of transposon-tagged plant genes. Reversion was shown to occur somatically, and its rate was temperature dependent. Inheritance studies showed that the mutations in two of the susceptible plants from the tagging protocol map at, or very close to, the race α-specific resistance gene. The results are consistent with the transposon tagging of a race-specific gene for rust resistance.     

Allelic composition and genetic background effects on transgene expression and inheritance in white clover

Allelic composition and genetic background effects on GUS expression and inheritance using a chimeric (cauliflower mosaic virus 35Sp:uidA) transgene were investigated in white clover as a prelude to transgenic cultivar development. Stable expression and Mendelian inheritance of the uidA transgene was observed over two generations when the uidA transgene was maintained in a heterozygous state. Transgenic backcross progeny (BC1) were intercrossed to produce segregating F2 populations. GUS-positive F2 plants were test-crossed with a non-transgenic control plant to determine whether individuals were heterozygous or homozygous for the transgene. Both expected and distorted segregation ratios were observed. Distortion of the segregation ratio was not caused by transgene inactivation or rearrangement, but was influenced by genetic background. BC1, BC2 and F2 populations were found to have similar levels of uidA gene expression. Quantification of GUS expression from progeny of high and low GUS expressing plants indicate that it is possible to alter transgene expression through selection. No difference was found between the level of expression for F2 plants homozygous or heterozygous for the transgene. These results indicate that F2 plants, homozygous for a transgene, might be used to develop a transgenic cultivar. However, progeny testing to determine the influence of genetic background is a prerequisite to such a development.     

A Second Locus, Ixr B1 in Arabidopsis thaliana, that Confers Resistance to the Herbicide Isoxaben

An isoxaben resistant mutant of Arabidopsis thaliana is described whose locus, Ixr B1, is unlinked genetically to the previously described resistance locus Ixr A (DR Heim, JL Roberts, PD Pike, IM Larrinua [1989] Plant Physiol 90: 146-150). A cross of strains each homozygous for one of these two resistance loci gives rise to some isoxaben sensitive F₂ progeny. Growth curves versus isoxaben of this mutant, its F₁ progeny and the wild-type parent strain showed that this locus displays a weakly codominant Mendelian phenotype. Callus cultures were established from plants homozygous as well as heterozygous for this locus. Growth inhibition curves done with these cultures mimic the data obtained in vivo.     

Spontaneous and bleomycin-induced genomic alterations in the progeny of Drosophila treated males depends on the Msh2 status. DNA fingerprinting analysis

Deficiency in DNA mismatch repair (MMR) confers instability of simple repeated sequences and increases susceptibility to cancer. Some of the MMR genes are also implicated in other repair and cellular processes related to DNA damage response. Supposedly, lack of their function can lead to a global genomic instability, besides microsatellite instability (MSI). To study the spontaneous and induced genomic instability in germ cells, related to the Msh2 status, DNA alterations in the progeny of individual crosses of Drosophila deficient in one or two copies of the Msh2 gene, were analysed by the arbitrarily primed polymerase chain reaction (AP-PCR). The results indicate that the progeny of homozygous parents for the normal Msh2 allele (+/+) presents a significantly lower frequency of genomic alterations than those from heterozygous (+/-) or mutant homozygous (-/-) parents. In addition, the DNA damage transmitted to the progeny, after the adult parental males were exposed to bleomycin, indicates that whereas the induction of mutations related to MSI depends on the lack of the Msh2 function, the induction of other mutational events may require at least one functional Msh2 allele. Thus, the results obtained with heterozygous individuals may have special relevance for cancer development since they show that a disrupted Msh2 allele is enough to generate genomic instability in germ cells, increasing the genomic damage in the progeny of heterozygous individuals. This effect is enhanced by mutagenic stress, such as occurs after bleomycin exposure.     

Genetic analysis of androgenetic rainbow trout.

We analyzed a number of genetic characteristics in androgenetic rainbow trout (Oncorhynchus mykiss) and their progeny. The androgenetic progeny of individual androgenetic males appeared genetically identical to each other based on eight enzyme loci. Their viability was no higher than that of androgenetic progeny of outbred males. Homozygous androgenetic female rainbow trout produced very poor quality eggs. When common eggs and sperm from outbred individuals were used to produce androgenetic and gynogenetic progeny, the yield of gynogenetic progeny was higher but some were heterozygous at protein loci, while no androgenetic progeny were heterozygous. Some androgenetic diploid rainbow trout were successfully produced from cryopreserved sperm. The progeny of some androgenetic males crossed to normal females were virtually all males, while the progeny of other males were virtually all females. This suggests that both XX and YY androgenetic individuals may develop as males. Androgenesis is likely to be useful for generating homozygous clones for research and for recovering strains from cryopreserved sperm.     

The influence of the genes An1, An2, and An4 on the activity of the enzyme UDP-glucose:Flavonoid 3-O-glucosyltransferase in flowers of Petunia hybrida

A relation between gene dosage and UDP-glucose:flavonoid 3-O-glucosyl-transferase (UFGT) activity was found in homozygous dominant and recessive parental lines and their F₁ progeny for both of the genes An1 and An2. In both F₂ crosses, progeny plants could be classified as belonging to groups showing either a low or a medium to high UFGT activity. Test crosses showed that heterozygous and homozygous dominant plants were present throughout the medium- to high-active group. The dosage relation in F₂ plants is most probably confounded by the segregation of modifiers. Thermal inactivation experiments indicated that structurally different UFGT enzymes are formed in homozygous dominant lines as well as in lines homozygous recessive for either An1 or An2. Lines homozygous recessive for the gene An4 contain a UFGT with a half-life time at 55° C of less than 8 min, whereas UFGTs from lines homozygous dominant for An4 show a half-life time of 25 min or above, with one exception. This relation was confirmed in the F₂ progeny; heterozygotes for An4 showed an intermediate half-life time. It is concluded that An4 might be the structural gene for the enzyme; An1 and An2 are both regulatory genes. UFGT activity in flowerbuds of An4/An4 plants seems to be lower than in an4/an4 plants. Anthers of flowers of an4/an4 lines, however, are virtually devoid of UFGT activity.     

S-related protein can be recombined with self-compatibility in interspecific derivatives ofLycopersicon

Stylar proteins involved in the self-incompatible (SI) response ofLycopersicon hirsutum have been identified and mapped to the locus that controls SI (S locus).L. esculentum, a self-compatible (SC) species of cultivated tomato, does not display these proteins. Hybrids between SCL. esculentum and SIL. hirsutum are self-sterile despite these individuals bearing pollen containing theS allele ofL. esculentum. In progeny derived from backcrossing the hybrids toL. esculentum, there was a strong correlation between the presence of theS allele fromL. hirsutum and self-infertility. However, this relationship was uncoupled in a number of backcross (BC) progeny. The SI response appeared to be nonexistent in two self-fertile BC individuals that were heterozygous for theS allele ofL. hirsutum, based on Mendelian segregation of a tightly linked DNA marker,CD15, in selfed progeny. Among these progeny self-fertile individuals that were homozygous for theL. hirsutum allele of the linked marker were also determined to be homozygous for anS-related protein ofL. hirsutum through test crosses withL. esculentum. Therefore, plants were produced that were homozygous for a functionalS allele but were self-fertile. This result and other evidence suggest that theS-related proteins are not sufficient to elicit a self-incompatible response inL. esculentum and that there is a mutation(s) inL. esculentum somewhere other than theS locus that leads to self-compatibility.     

Suppression and Function of X-Linked Lethal and Sterile Mutations in CAENORHABDITIS ELEGANS

We have expanded our collection of recessive lethal and sterile mutants in the region of the X chromosome balanced by mnDp1(X;V), about 15% of the X linkage map, to a total of 54 mutants. The mutations have been mapped with respect to 20 overlapping deficiencies and five X duplications, and they have been assigned to 24 genes by complementation testing. Nine mutants are hermaphrodite-sterile: one of these is a sperm-defect mutant, two have abnormal gonadogeneses and six, in five genes, are maternally influenced mutants, producing inviable zygote progeny. One of the gonadogenesis mutants and two of the maternally influenced mutants are male fertile. All but one of the maternally influenced mutants give cross progeny when mated with wild-type males. Forty-three mutants were tested for suppression by homozygous sup-5(e1464), which is believed to be specific for null alleles. Ten mutants that were judged by independent criteria not to be null mutants are not suppressed. Nine of the other 33 mutants, in nine genes, are suppressed, five in both heterozygous and homozygous suppressor stocks and four only in homozygous suppressor stocks.     

The genetic interaction between non-nodulation and supernodulation in soybean: an example of developmental epistasis

Summary The interaction between three non-nodulation mutants (nod49, nod772 and nod139) and a supernodulation mutant (nts382) of soybean was studied by analysing the progeny from crosses between these mutants. Previously it had been shown that the non-nodulation mutants arose from single mutation events and that nod49 and nod772 are allelic, whereas nod139 represents another gene required for nodulation. Analysis of progeny from crosses between nts382 and the wild type showed that this mutant also arose from a single mutation. Complementation tests demonstrated that the mutation responsible for supernodulation in nts382 is not allelic to either of these non-nodulation characters, and that it segregates independently. Progeny were identified that were homozygous for both supernodulation and non-nodulation, and these plants were incapable of nodulation. Thus, non-nodulation is epistatic over supernodulation and this is discussed in terms of the developmental blockage in the two mutant types. The identification and confirmation of these double mutants of the supernodulation and non-nodulation mutations are described. Although the non-nodulation mutations behave as recessive characters in a wild-type background, these mutations are incompletely dominant in a genetic background homozygous for supernodulation. The significance of these results to the understanding of nodule ontogeny is discussed.     

Combining transgenic and natural resistance to obtain broad resistance to tospovirus infection in tomato (Lycopersicon esculentum mill)

This study was undertaken to develop tomato plants with broad resistanceto tospoviruses which are a major limiting factor to tomato productionworldwide. A nontransgenic tomato line Stevens-Rodale (S-R), six transgenictomato lines expressing the nucleocapsid (N) protein gene of the lettuceisolate of tomato spotted wilt virus (TSWV-BL), and progeny of the crosses between S-Rand three of the transgenic lines homozygous for the N gene were evaluated fortheir resistance to tospovirus infection in greenhouse inoculation tests. S-Rhas the Sw-5 gene that confers resistance to several TSWVisolates. The six transgenic lines showed high levels of resistance wheninoculated with either TSWV-BL or a tomato isolate from Hawaii (TSWV-H).However, these same plants were highly susceptible to the Brazilian isolate ofgroundnut ringspot virus (GRSV-BR). Plants with the Sw-5gene were resistant to TSWV-BL and GRSV-BR, but were susceptible to TSWV-H.When inoculated with any of the three viruses, the F₁ progeny of thecrosses exhibited a susceptible, tolerant, or resistant phenotype with a higherproportion of the plants being either tolerant or resistant. When F₂progeny from F₁ resistant plants of each cross were inoculated withany of the three viruses, a higher proportion of tolerant and resistant plantswas observed compared to the F₁ progeny. Our results show thepotential to obtain broad resistance to tospoviruses by combining transgenicand natural resistance in a single plant.     

Failure of X-rays to mutate class II histocompatibility loci in balb/c mouse spermatogonia

Adult Balb/c Kh male mice were irradiated (pelvic region, 250 kVcp X-rays, 60 rad per min) and three months later were mated to untreated C57BL/6 Kh females. Their B6C F₁ progeny were screened for mutations at the Class II histocompatibility loci, i.e. those that carry similar alleles in the parental lines and are therefore homozygous in the F₁ progeny. The treatment groups were: single doses of 0, 350, 500, 650 and 800 rad; split doses 1 day apart, totalling 500, 650 and 800 rat; split doses averaging 52 days apart, totalling 650 and 800 rad. Thirty-six mutants were identified in 13,614 progeny. Twelve of them occurred in five clusters of two or three, presumably owing to five gonadal mosaics among 940 parents. Irradiation did not increase the spermatogonial mutation rate. The only effect of exposure appeared to be a decrease in the mutation rate of the 1-day split dose-groups compared to those with the same total doses in a single exposure or in two fractions, 52 days apart.