Interstrain Crosses Research Articles

A Y-chromosomal effect on blastocyst cell number in mice.

Karyotopic and cell number analysis of 3.5 day post coitum preimplantation mouse embryos was used to determine whether XY embryos had more cells than XX embryos at the late morula/early blastocyst stage. This proved to be the case for the CD1 strain (for which it had previously been shown that XY embryos form a blastocoel earlier than XX embryos) and for the MF1 strain. However, this increased cell number was not seen in MF1 embryos carrying an RIII strain Y in place of the MF1 Y. Furthermore, interstrain crosses between CD1 and the MF1,YRIII strain showed that the cell number increase segregated with the CD1 Y but not with the RIII Y. It is concluded that the CD1 and MF1 Y chromosomes carry a factor that accelerates the rate of preimplantation development.

Generation of a restriction fragment length polymorphism linkage map for Toxoplasma gondii.

We have constructed a genetic linkage map for the parasitic protozoan, Toxoplasma gondii, using randomly selected low copy number DNA markers that define restriction fragment length polymorphisms (RFLPs). The inheritance patterns of 64 RFLP markers and two phenotypic markers were analyzed among 19 recombinant haploid progeny selected from two parallel genetic crosses between PLK and CEP strains. In these first successful interstrain crosses, these RFLP markers segregated into 11 distinct genetic linkage groups that showed close correlation with physical linkage groups previously defined by molecular karyotype. Separate linkage maps, constructed for each of the 11 chromosomes, indicated recombination frequencies range from approximately 100 to 300 kb per centimorgan. Preliminary linkage assignments were made for the loci regulating sinefungin resistance (snf-1) on chromosome IX and adenine arabinoside (ara-1) on chromosome V by linkage to RFLP markers. Despite random segregation of separate chromosomes, the majority of chromosomes failed to demonstrate internal recombination events and in 3/19 recombinant progeny no intramolecular recombination events were detected. The relatively low rate of intrachromosomal recombination predicts that tight linkage for unknown genes can be established with a relatively small set of markers. This genetic linkage map should prove useful in mapping genes that regulate drug resistance and other biological phenotypes in this important opportunistic pathogen.

A genetic mapping system in Caenorhabditis elegans based on polymorphic sequence-tagged sites.

We devised an efficient genetic mapping system in the nematode Caenorhabditis elegans which is based upon the differences in number and location of the transposable element Tc1 between the Bristol and Bergerac strains. Using the nearly completed physical map of the C. elegans genome, we selected 40 widely distributed sites which contain a Tc1 element in the Bergerac strain, but not in the Bristol strain. For each site a polymerase chain reaction assay was designed that can distinguish between the Bergerac Tc1-containing site and the Bristol "empty" site. By combining appropriate assays in a single reaction, one can score multiple sites within single worms. This permits a mutation to be rapidly mapped, first to a linkage group and then to a chromosomal subregion, through analysis of only a small number of progeny from a single interstrain cross.

Stimulation of Oviposition and Heterosis in Interstrain Crosses of European Corn Borer (Lepidoptera: Pyralidae)

Hybrid matings of two Ostrinia nubilalis (Hübner) (Lepidoptera: Pyralidae) colonies (N and O), established in the laboratory a year apart, demonstrated positive heterosis in the F1 generation as indicated by increases in pupal weight (6%), number of egg masses per female (65%) and number of eggs per female (77%) as compared to intra-colony crosses in mass matings. In contrast, pupation date, an indicator of development time, was intermediate in the hybrids relative to the parental colonies. There was also a 37% increase in the numbers of eggs/female and a 45% increase in egg masses/female in the hybrid parental cross. We investigated this stimulation of oviposition in single female, reciprocal crosses. Oviposition stimulation was directional with the cross of N colony males x O colony females exhibiting significantly higher numbers of total eggs/female and egg masses/female on a daily basis. Colony O females in this cross laid more large egg masses than females in other crosses. Data from individual female crosses demonstrated that egg production in hybrid mass mating experiments was consistent with random mating of males and females from both colonies.

The identification of biological species in the genus Heterorhabditis (Nematoda: Heterorhabditidae) by cross-breeding second-generation amphimictic adults

Entomopathogenic nematodes of the genus Heterorhabditis are morphologically conservative, consequently the majority of isolates remain unassigned at the species level. When a Heterorhabditis infective juvenile infects an insect host, it develops into a protandrous hermaphrodite female. These first-generation females give rise to a second generation which contains both males and females. Because of this complex life-cycle and also because of uncertainty as to whether second-generation females are amphimictic, cross-breeding studies to facilitate species determination have not been carried out previously. We demonstrate here that second-generation Heterorhabditis females are amphimictic. Because of this finding, we have been able to develop a successful cross-breeding technique for the purposes of species determination in Heterorhabditis. Interstrain crosses using second-generation males and females from the appropriate strains have been successfully set up in Xenorhabdus luminescens-treated G. mellonella cadavers and also on agar plates. Using the techniques described here we confirm that H. bacteriophora (Brecon strain), H. megidis and H. zealandica are distinct biological species, we note that the H. bacteriophora group contains at least 2 species and we provide evidence for the existence of a new Irish species of Heterorhabditis.

Stability of Dicofol Resistance in Populations of European Red Mite (Acari: Tetranychidae) on Apples in Southern Ontario

Fluctuations in levels of resistance to dicofol in populations of European red mite, Panonychus ulmi (Koch), from several orchards with different frequencies of resistant individuals were compared with responses of laboratory colonies established from the same sites. Considerable variability in responses occurred in observations spaeed over 2 yr, but the frequency of dicofol-resistant individuals in these populations did not decline in the absence of selection pressure. Laboratory colonies reared through 32 generations displayed relatively constant resistance frequencies, suggesting that dicofol resistance is stable for a time equivalent to at least 3 yr in the field. Relative fecundities of mites resistant and susceptible to dicofol were examined by comparing the number of eggs laid by females from intra- and interstrain crosses, as well as the percentage hatchability, percentage survivorship, and sex ratio of the resulting progeny. Females from a 12-yr-old susceptible laboratory colony laid more eggs than females from a more recently established susceptible colony and than females from two colonies resistant to dicofoi. Resistant and susceptible mites did not differ for any of the other parameters examined. It is concluded that the absence of a reproductive detriment is a major factor in the apparent stability of resistance to dicofol in these mites. Implications of these findings for management of dicofol resistance in P. ulmi are discussed.

Reproductive compatibility and mitochondrial DNA restriction site analysis of New World screwworm, Cochliomyia hominivorax, from North Africa and Central America

The reproductive compatibility of New World screwworms, Cochliomyia hominivorax (Coquerel), from North Africa and a strain being mass produced for the Mexican eradication programme was examined to assess the feasibility of using flies from the Mexican screwworm mass production facility for a sterile insect technique eradication programme in North Africa. Males from the production strain mated randomly with females from North Africa and from the production strain when both were present. Neither strain of males discriminated between cuticular extracts of North African and production strain females containing a contact sex pheromone. Interstrain crosses between North African flies and production flies were fertile and produced fertile progeny. Chromosome morphology did not differ significantly between the two strains and homologue pairing was normal in hybrid meiotic and polytene nuclei. Mitochondrial DNA restriction site analyses indicated that the genetic divergence of the North African strain from Mexican and Central American strains was within the range of the diversity observed in Central American, Mexican and Caribbean populations. Test results indicate that New World screwworms from North Africa are reproductively compatible with the strain currently being mass produced in Mexico. Mating barriers should not impede the progress of an eradication programme using the sterile insect technique in North Africa with sterile screwworms from the Mexican mass production facility.

Growth and Survival of Intrastrain and Interstrain Rainbow Trout (Oncorhynchus mykiss) Triploids1

AbstractTriploid rainbow trout exhibit improved survival and extended growth during sexual maturation, compared to their diploid counterparts. However, there have been few benefits demonstrated prior to sexual maturation. This study was undertaken to investigate the possibility of improving growth and survival parameters in triploids through interstrain crosses. Triploids were induced by heat shocking fertilized eggs from intra‐ and interstrain crosses of two rainbow trout strains. The four triploid groups and their diploid counterpart groups were reared to 233 days post‐hatching and analyzed for growth and survival characteristics.Compared with diploids, triploids had significantly (P < 0.05) higher mortalities during the first 100 days post‐fertilization, primarily just prior to and after hatching. However, during the remainder of the study triploids exhibited significantly (P < 0.05) lower mortalities than diploids. During the first 50 days of rearing all four triploid groups were significantly shorter and lighter than their diploid counterparts. The growth of the triploid groups later in the study varied considerably. At the conclusion of the rearing phase, one interstrain triploid group was significantly (P < 0.05) longer than its diploid counterpart, although not significantly heavier. The other triploid groups were either significantly smaller than, or equal to the diploids. Analysis of variance indicated that the growth of triploid rainbow trout was significantly affected by maternal strain effects. These results suggest that the use of specific strains and crosses may improve the growth of triploid rainbow trout.

Genetic Evidence for Sibling Species within the Sugarcane Borer (Lepidoptera: Pyralidae)

Collections of the sugarcane borer, Diatraea saccharalis (F.), from Brazil and Louisiana differ in morphology, physiology, and behavior. These samples, along with one from Mexico, were examined electrophoretically to aid in determining the basis of this variation. Two other species, D. grandiosella Dyar and D. lineolata (Walker), were also examined. Genetic differences were minor between Mexican and Louisiana samples (Nei's distance = 0.02) and an order of magnitude greater in pairwise comparisons with the Brazilian sample (average distance = 0.23). The Brazilian collection was significantly different at all polymorphic loci, and one locus was diagnostic. Although D. grandiosella and D. lineolata are nearly identical morphologically, they differed greatly genetically (Nei's distance = 0.36; three diagnostic loci). Variation within the sugarcane borer is well within the range of interspecific differences, suggesting D. saccharalis may be composed of two species. Interstrain crosses performed using Louisiana and Brazilian laboratory colonies indicated no incompatibilities in parental, interhybrid, or backcrosses, suggesting postreproductive barriers may not exist. Further studies are required to determine the geographical ranges of the two taxa, and, if sympatric, whether they are completely reproductively isolated in nature.

Intestinal leucine aminopeptidase and alkaline phosphatase: Genetic regulation and development in mice

Intestinal and serum leucine aminopeptidase (LAP) and alkaline phosphatase (AKP) were characterized by electrophoresis for eight inbred strains of laboratory mice. Intestinal LAP and AKP of adult mice were expressed concordantly within strains, as banded or diffuse, and concordantly for rate of migration within strains that had diffuse isozymes. All strains, except DD/S, had a single band of serum LAP and a single, diffuse zone of serum AKP. DD/S had a double band of serum LAP as well as isozymes of intestinal LAP and AKP unlike those of other strains. All strains displayed similar, neuraminidase-sensitive isozymes of intestinal LAP and of AKP prior to weaning, but after weaning there was marked sensitivity to neuraminidase only in DD/S. In interstrain crosses, banded/diffuse, migration rate, and neuraminidase sensitivity were inherited as independent autosomal traits, with indications of variable penetrance and genetic interaction.

Tissue and strain-specific patterns of endogenous proviral hypomethylation analyzed by two-dimensional gel electrophoresis.

Proviral sequences related to the intracisternal A particle (IAP) are amplified and dispersed in the mouse genome. Their expression is associated with hypomethylation at CpG sites in the 5' long terminal repeat. We have used two-dimensional agarose gel electrophoresis to examine patterns of IAP hypomethylation in mouse DNA. The method is sensitive to both the methylation status of a conserved Hae II site in the 5' long terminal repeat and the location of the closest BamHI site in the flanking DNA upstream of each hypomethylated long terminal repeat. The method also defects restriction fragments derived from IAP elements that are themselves methylated but have an unmethylated Hae II site in their 5' adjacent DNA. DNAs from each of four inbred mouse strains (BALB/c, C3H/He, C57BL/6, and DBA/2) gave distinctive two-dimensional patterns of BamHI/Hae II restriction fragments detected by hybridization with an IAP probe. This constitutive pattern was largely conserved among several tissues of each strain, but some tissue-specific variations were observed. The site-specific hypomethylations reflected in the two-dimensional patterns were heritable properties, since DNA from progeny of an interstrain cross contained both parental sets of fragments. IAP elements may be useful endogenous reporters of genomic methylation patterns.

Inheritance of two endosperm protein loci in rice (Oryza sativa L.)

Previous studies indicated two types of phenotypic protein markers as two minor bands of SDS-PAGE for rice storage protein. A variant derived from a Pakistani variety, Dular, was found to show a mobility variant with Band 11, a relatively faster-moving band as compared to Band 10, while most of the other cultivated rices exhibited Band 10 at a molecular weight of around 100-110 K. Band 11 was also observed in several wild rice species. How this variant occurred is not known. Another marker is characterized by the presence of either Band 56 (slower-migrating band) or Band 57 (faster-migrating band) in most cultivars at a molecular weight of about 28-27 K. Most indica varieties developed in Taiwan have Band 57 and japonica varieties have Band 56. Genetic analysis of F1, F2 and F3 seeds from interstrain crosses indicated that Band 10 versus Band 11 and Band 56 versus Band 57 are due to codominant alleles at two loci. Tests of independent inheritance between these two loci (Band 10/11 versus Band 56/57) indicated that there is no linkage between them. Both of these two protein loci encode for endosperm proteins and mostly belong to the minor polypeptide subunits of the glutelin fraction of rice seed proteins. Studies on reciprocal crosses indicate dosage effects as exhibited in band patterns. Variations in band intensity were frequently observed when the maternal genotype was different.

The Tc2 transposon in Caenorhabditis elegans.

A second family of transposons, named Tc2 elements, has been identified in the nematode Caenorhabditis elegans. Tc2 elements are polymorphic in sequence and are present in different numbers in different strains. Like the transposon Tc1, Tc2 is active in the germ line of some C. elegans strains. A high rate of transposition has been observed in the progeny of certain interstrain crosses, where transposition events are frequent enough to be detected in blot hybridization experiments, without the use of a genetic screen. Our data suggest that transposition of Tc1 and Tc2 may be regulated by the same genomic factors.

Detection of a null allele for MDH isozymes in the guppy (Poecilia reticulata), with special reference to sex-linked inheritance.

Malate dehydrogenase (MDH) was examined by starch gel electrophoresis for strain differences in the guppy, Poecilia reticulata. The electrophoretic patterns revealed duplicate gene loci encoding a mitochondrial form of MDH and the existence of a null allele at one of the duplicate loci. Different phenotypic frequencies at Mdh-4 were observed among 9 strains and between males and females within each strain. The progeny tests for Mdh-4 variants in the inter-strain crosses indicated a mode of partial sex-linked inheritance.

Reduced fertility in gracile axonal dystrophy (gad) mice.

Mean litter size in gad/gad females was significantly lower than in normal females (+/+ and gad/+) in intra- and inter-strain crosses. The reduction in litter size was not dependent on the genotypes of the males, but could be attributed to the gad/gad females themselves. The numbers of corpora lutea and implants in gad/gad females were slightly reduced as compared with those in the controls, but the number of live fetuses was significantly lower than that in normal females 14 days after copulation (P less than 0.02). Hence, reduced litter size in gad/gad females was accounted for mostly by embryonic and fetal death after implantation, which was inferred to be due to impaired uterine function.

MATING BETWEEN DIAPAUSING AND NONDIAPAUSING STRAINS OF THE CITRUS RED MITE, PANONYCHUS CITRI (MCGREGOR)

AbstractThe reproductive compatibility between diapausing (DP) and nondiapausing (C) strains of the citrus red mite, Panonychus citri (M.), was determined by crossing experiments. Mating between the two strains was unrestricted, but copulation between C females and DP males was often broken off prematurely, whereas copulation in the reciprocal cross was of normal duration. Diapausing males showed a strong preference for guarding quiescent female deutonymphs of their strain prior to copulation, whereas C males did not. However, males of either strain showed no preference between adult females, resulting in only a partial, one-sided premating isolation between strains. No adult females were produced in the progeny from either interstrain cross, showing reproductive incompatibility is complete. Hatchability of the eggs from the cross between DP females and C males was significantly lower than that from the cross between C females and DP males, or from the intrastrain crosses, suggesting that even though some of the eggs in the former cross were fertilized, they died during embryonic stages. This possibility was supported by the observation that, for DP females mated with a C male, the second intrastrain mating was almost totally ineffective. Also, for C females mated with a DP male, a second intrastrain mating was often ineffective, but to a lesser extent than for DP females. When the two strains coexisted, effective mating was severely curtailed for both strains. The results explained the low proportion of females from overwintered DP eggs in a pear orchard in which the two strains coexisted the previous autumn.

Life cycle, meiotic recombination and unilateral gene transfer in methanolotrophic yeasts of the genus Pichia

In order to perform strain improvement of the methanol utilising yeasts Pichia pinus and Pichia lindneri their life cycles were elucidated. Both yeasts are monoecious and self-fertile. Karyogamy by autodiploidization is followed by meiosis leading to Mendelian segregation within the offspring. In Pichia pinus crosses via cell or protoplast fusion is feasible as proven by tetrad analysis. This is not the case in Pichia lindneri. The only possiblity to obtain recombinants in this yeast is brought about by a yet unknown mechanism, leading to a unilateral transfer of genetic material, found after cocultivation of mutants. This is revealed by the evaluation of F1 offspring from meiotic divisions after autodiploidisation. Although analysis of nuclear and mitochondrial DNA revealed rather gross differences not only between the species but also within different strains of Pichia pinus, only a limited exchange of genetic material is possible in interstrain crosses.

Absence of Diapause in a Laboratory Strain of the Florida Viceroy Butterfly

Larvae of the southern subspecies Limenitis archippus floridensis were reared under eight photoperiod regimes (varying from LD 8:16 to LD 18:6) to determine the diapause threshold (D50; the daylength which induces 50% diapause) and critical range of photoperiods over which the frequency of diapause changes from low to very high. Previous determinations based on Vermont and Maryland strains of L. archippus archippus suggested by extrapolation thatfloridensis larvae should have a critical range between LD 10.5:13.5 and LD 12.5:11.5, with a D50 of 11.5 hr. However, all 220floridensis larvae reared at room temperature developed directly to fourth instar without diapause, regardless of photoperiod. Four interstrain crosses (fzoridensis females x Maryland archippus males) reared under LD 8:16 photophase varied widely in diapause (0-94 7O), suggesting that facultative larval diapause in this species is under polygenic control. Reasons for the absence of diapause in this floridensis strain are suggested. INTRODUCTION Many insects alter their growth and behavioral activities in response to seasonal changes. Before the onset of cold weather and short food supplies, insects in temperate climates must either diapause or migrate to a less stressful environment. Various ecological factors have been postulated to induce diapause. Photoperiod is considered to be the most important cue because it correlates well with the seasons and does not fluctuate as much as other environmental variables (Beck, 1980; Danilevskii et al., 1970). There is evidence in some species that warm temperature interacts with photoperiod and reduces the incidence of diapause (Adkisson et al., 1963; Beck, 1977; Beach, 1978; Caldwell and Wright, 1978; Hayes, 1982). The condition of the food plant affects diapause onset in some species (Danilevskii et al., 1970; Dingle et al., 1977). The ability to diapause contributes to the evolutionary success of many temperate species (Chippendale, 1982). Insects with wide geographic distributions vary in photoperiod responses. Specifically, long-day insects (which diapause in the late summer and early autumn) do so in response to longer photoperiods in northern populations and shorter photoperiods in southern populations (Beck, 1980; Danilevskii et al., 1970). This variation in photoperiod threshold has been reported by Beck (1963) to occur in Ostrinia nubilalis (Hiibner) (Lepidoptera: Pyralidae), by Tauber and Tauber (1972) in Chrysopa carnea Stephens (Neuroptera: Chrysopidae), by Masaki (1972) in Pteronemobius fascipes Walker (Orthoptera: Gryllidae), by Sims (1982) in Aedes triseriatus (Say) (Diptera: Culicidae), and by Hong and Platt (1975) in Limenitis (Basilarchia) archippus (Cramer) (Lepidoptera: Nymphalidae). The viceroy butterfly (Limenitis archippus) is common and broadly distributed throughout North America, ranging from southern Canada to Mexico. It prefers moist meadows and disturbed localities in the primary stages of succession, where its food plants, shrub willows and poplars (Salicaceae) establish rapidly. During late summer and autumn in temperate climates the third instar larvae exhibit facultative diapause in response to short-day photoperiod. They construct hibernacula in the basal portions of tubular willow leaves covered with silk (Scudder, 1889; Weed, 1926; Klots, 1951; Frankos and Platt, 1976). These tubular chambers remain open at their distal ends. This overwintering behavior is characteristic of all temperate species of the Limenitini (Lederer, 1960).

Protoplast fusion and genetic recombination in Metarhizium anisopliae

Using two strains of Metarhizium anisopliae var. minor designated E6 and RJ from different origins, inter-strain crosses were readily performed by orthodox methods from mutants of strain E6 through the parasexual cycle. However, in most cases crosses between mutants of strain RJ or between RJ and E6 mutant strains were not achieved. Protoplast fusion was carried out in an attempt to cross such strains. Protoplasts were obtained after treatment of mycelium with lytic enzymes using 0.7 m KCl as osmotic stabilizer. Regeneration frequency varied from 1.2 to 3.0%. Poly (tethylene glycol) was used for the fusion of protoplasts and fusion frequencies varied from 0.9 to 44.1 in 10 5 protoplasts according to the cross. Sectors which emerge from fusion products were analysed and recombinants were obtained even from crosses between mutant strains which could not be crossed by hyphal fusion. In this way protoplast fusion proved to be a valuable tool for further studies of genetics and breeding of Metarhizium anisopliae.

Cryopreservation of strains and mutant genes in mice.

Three kinds of freezing methods were tested with embryos of DNI strain. The survival rate after thawing was 47.5%, 66.7% and 77.8% in the 2-step method, modified slow freezing method and modified 2-step method, respectively. Then, the modified 2-step method was applied to the embryos from 7 strains and a pair of interstrain crosses. PMSG treatment at the beginning of diestrus following HCG treatment after 48 hrs resulted in much yield of 8-16-cell embryos in all strains. The average number for each strain was as follows: DNI; 18.9, DDN; 13.0, BS; 20.4, C57BL/6; 12.9, DBA/2; 17.5, CRN; 19.8, PAN; 13.7 and DNI x C57BL/6-Ay; 21.7. Development of frozen-thawed embryos in culture varied among strains. Proportion of embryos that developed to the morula or blastocyst stage was as follows: DNI; 64.6%, DDN; 71.9%, BS; 53.6%, C57BL/6; 57.3%, DBA/2; 65.0%, CRN; 52.5%, PAN; 17.4% and DNI x C57BL/6-Ay; 44.1%. These results indicate that the ability of embryos to survive freezing and thawing is influenced by their genetic background. Live young were produced from DNI, DDN, BS and DNI x C57BL/6-Ay embryos after transfer to recipients. Comparative assessment of the developmental ability of frozen-thawed embryos after transfer among strains should be performed in further study.