Meiotic Chromosome Preparations Research Articles

Effects of the DNA topoisomerase II inhibitor merbarone in male mouse meiotic divisions in vivo: cell cycle arrest and induction of aneuploidy.

In order to clarify possible risks of aneuploidy induction in germ cells by cancer chemotherapy we studied effects of a non complex-stabilizing DNA topoisomerase II (topo II) inhibitor merbarone in male mouse meiotic divisions in vivo. Two cytogenetic approaches were used: (1) C-banding on meiotic chromosome preparations and (2) analysis of spermatid micronuclei (MN) combined with immunocytochemical staining of kinetochore proteins using CREST serum. For comparison, another topo II inhibitor, VP-16, known to form cleavable complexes, was studied. The microdissection technique of mouse seminiferous tubules enabled us to carefully examine effects at specific phases of meiosis. Merbarone injections increased percentages of polyploid and hypoploid metaphase II spermatocytes at time intervals corresponding to the treatment of the first meiotic division and diplotene-diakinesis. The highest level of MN induction (5.8 MN/1000 spermatids, P < 0.001) was observed in animals injected 48 hours before the harvest, corresponding to the treatment of diplotene-diakinesis spermatocytes. Most of the induced MN (80%) contained kinetochore signals, indicating that they resulted from detachment of a whole bivalent or chromosome from the meiotic spindle. The high frequency of MN with two kinetochore signals at opposite sides (33%) most likely denotes lagging of whole bivalents during MI. Inhibition of cell proliferation was determined by scoring cells arrested at different phases of MI and MII. All groups of treated animals showed a clear increase in the frequency of arrested divisions compared to controls (P < 0.001). Thus, merbarone was shown to severely damage normal meiotic processes.

Chromosomes of three prosobranch gastropods from Viviparidae, Pilidae and Cyclophoridae (Order: Mesogastropoda)

SUMMARYMitotic and meiotic chromosome preparations of Bellamya bengalensis f. doliaris (Viviparidae), Pila globosa (Pilidae) and Cyclophorus polynema (Cyclophoridae) were made by colchicine—ovotestis cell suspension—methanol acetic acid—flame drying—Giemsa technique. Karyotypes were prepared on the basis of decreasing length of the chromosomes and their morphometric analyses were done. The diploid number 2n = 22 with the chromosome formula n = 7m + 4 sm and FN = 44 for Bellamya bengalensis f. doliaris, 2n = 28 with the chromosome formula n = 9m + 5sm and FN = 56 for Pila globosa and 2n = 28 with the chromosome formula n = 13m + 1sm and FN = 56 for Cyclophorus polynema were obtained. Chromosome numbers of each species was confirmed from the respective metaphase-I spreads and behaviour of their meiotic chromosomes were studied. The data obtained for each species were compared with the available chromosomal information on other confamilial species and the role of the possible mechanisms of karyotypic evolution in them have been discussed. Conservativeness with regard to chromosomal change at higher taxon level has been discussed.

Chromosomal location of the amplified esterase genes conferring resistance to insecticides in Myzus persicae (Homoptera: Aphididae)

A genomic probe encompassing most of an esterase gene (E4) that is amplified in insecticide-resistant Myzus persicae was hybridized in situ to mitotic and meiotic chromosome preparations of aphid clones of known esterase type and resistance level. Binding, which was detected using the biotin-avidin system located both known types of amplified esterase sequences (E4 and FE4). All except one of the E4-producing clones had a single amplified site, on autosome 3 near the breakpoint of an autosomal 1,3 translocation which previous work had shown to be genetically linked to insecticide resistance. The exceptional clone had two other E4-encoding sites. The most resistant FE4-producing clone (800F) had amplified sequences at five sites (three loci: two homozygous and one heterozygous). Altogether, amplified E4 and/or FE4 sequences were found on four of the five autosome pairs of M. persicae. Possible origins of these multiple loci are discussed.

A reciprocal whole-arm translocation, rcp(1;6)(1p6p;1q6q) in a boar, localization of the breakpoints, and reassignment of the genes for glucose phosphate isomerase (GPI) and calcium release channel (CRC).

A reciprocal whole-arm translocation between chromosomes 1 and 6 in a Swiss Large White boar with reduced fertility was identified by the use of different staining techniques in mitotic metaphase cells, synaptonemal complex analyses, and meiotic chromosome preparations. The karyotype of this boar was demonstrated to be 38,XY,rcp(1;6)(1p6p;1q6q). To further localize the breakpoints more precisely and determine the precise gene locations, several in situ hybridization experiments were performed with a chromosome 1 centromere-specific probe and two other gene probes. The breakage and reunion points of both chromosomes were located in the centromeric regions. The genes for glucose phosphate isomerase and calcium release channel were mapped to 6cen----q12.

Organization of a cloned repetitive DNA fragment in mosquito genomes (Diptera: Culicidae)

The structure and genomic organization of a cloned 5.2-kb repetitive DNA fragment, H-85, isolated from the Aedes albopictus genome have been examined. In situ hybridization of the 3H-labeled H-85 DNA to the meiotic and mitotic chromosome preparations of Ae. albopictus shows that the sequences homologous to H-85 DNA are dispersed throughout the length of all three pairs of chromosomes. A similar pattern of in situ hybridization appears in Aedes seatoi, Aedes flavopictus, and Aedes aegypti. The study shows that the arrangement of sequences in the cloned 5.2-kb fragment is rare in the Ae. albopictus genome. Dot-blot hybridization reveals that the sequences homologous to H-85 DNA are present in 12 species of mosquitoes examined, belonging to six genera in subfamilies Culicinae ad Anophelinae. The H-85 sequences are also present in the genome of Mochlonyx velutinus of the nematocerous family Chaoboridae, earlier proposed as the ancestor of the mosquito family Culicidae. Although the sequences homologous to H-85 DNA are present in different species of mosquitoes, they have diverged in their structure and organization. The cloned 5.2-kb fragment is composed of elements of different and independently evolving repetitive DNA families.

Localization of silver positive structures in mitotic and meiotic chromosomes ofKalotermes flavicollis(Fabr.) (Isoptera Kalotermitidae)

Application of the silver staining technique to mitotic and meiotic chromosome preparations of Kalotermes flavicollis have shown: (1) four pairs of mitotic chromosomes have nucleolus organizer regions located near the centromere regions; (2) that in suitable preparations core-like structures are visible in mitotic chromosomes; (3) the sex determining mechanism.

The chromosomes of tufted deer &lt;i&gt;(Elaphodus cephalophus&lt;/i&gt;&lt;i&gt;)&lt;/i&gt;

Mitotic and meiotic chromosome preparations of the tufted deer (Elaphodus cephalophus) were studied to elucidate the sex-chromosomal polymorphism evidenced by this species. Females had 2n = 46 or 47 chromosomes, whereas males had 2n = 47 or 48 chromosomes. An X;autosome translocation was identified by synaptonemal complex analysis of spermatocytes at pachytene and confirmed by the presence of a trivalent at diakinesis/metaphase I. The present work, in combination with earlier observations by others, indicates that E. cephalophus possesses a varied X-chromosome morphology involving an X;autosome translocation and addition of varying amounts of heterochromatin. It is speculated that sex-chromosome polymorphism may be responsible for the observed differences in diploid chromosome number of tufted deer.

Chromosomes of a pestiferous land snail,Achatina (Lissachatina) fulica fulica(Bowdich) (Achatinidae: Pulmonata: Gastropoda)

SUMMARYMitotic and meiotic chromosome preparations of Achatina (Lissachatina) fulica fulica were made by colchicine- citrate- flame drying- Giemsa technique. The diploid number 2n = 60 with the chromosome formula n = 28m + 2 sm and FN = 120 were obtained. On the basis of chromosome lengths the karyotype has been prepared and the morphometric analysis has been done. The haploid number n = 30 has been confirmed and the behaviour of germinal chromosomes have been studied. Conservativeness with regard to chromosomal change in Mollusca has been discussed.

Synaptonemal complex formation in male scorpions exhibiting achiasmate meiosis and structural heterozygosity

Synaptonemal complex (SC) formation was studied in testicular material from individuals of a number of species from two families of Australian scorpions; Buthidae and Scorpionidae. These scorpions exhibit unusual cytogenetic features including achiasmate male meiosis, interchange heterozygosity, and centromeric fusion–fission and inversion heterozygosity. The synaptic behaviour of chromosomes involved in these rearrangements was studied from zygotene to metaphase I, using both meiotic chromosome preparations and techniques for examination of the SCs. Multivalent associations present during the achiasmate meiosis of both buthid and scorpionid scorpions are retained from prophase to metaphase I, unlike those present in polyploid achiasmate Bombyx females. Further evidence suggests that synaptic adjustment does not occur generally in achiasmate scorpionid inversion heterozygotes. However, for some inversions, pairing is seen to become more heterosynaptic from late prophase to metaphase I and this may be related to the pairing maintenance system during achiasmate meiosis in these specialized organisms.Key words: synaptonemal complex, achiasmate meiosis, heterozygosity, interchange, inversion.

Meiosis in pollen mother cells of 15 Cryptomeria japonica clones with special emphasis on the behavior of the 6th chromosome pair heteromorphic to chromomycin A3 band.

Meioses in pollen mother cells (PMCs) were examined by the conventional staining and the CMA-staining methods in 15 clones of Cryptomeria japonica D. Don carrying three different karyotypes identified and classified by the characteristics of interstitial chromomycin A3 (CMA)-band or secondary constriction on the 6th chromosome pair. Before fixation of PMCs, a newly developed hypotonic treatment was applied for easy and constant preparation of well-spread meiotic chromosomes. At meiotic metaphase I 11 bivalents were observed, which possessed chiasmata at the interstitial region and/or the distal end. Although the CMA-bands were clearly observed on meiotic chromosomes, no chiasma was formed involving these bands. Most of the clones exhibited normal meiotic progression, but two clones (Syusou 5 and 11) showed some abnormal meioses. In Syusou 5 the heteromorphic 6th chromosome pair was involved in univalent formation more frequently than the other chromosome pairs. The normal meiosis in most clones with the heteromorphic 6th chromosome pair indicates that the heteromorphic chromosomes do not affect meiotic process.

Studies of the induction of dominant lethals and translocations in male mice after chronic exposure to microwave radiation.

Male C3H mice were exposed to 100 W m-2 of 2.45 GHz continuous-wave microwave radiation for 6 h per day for a total of 120 h over an 8-week period. The exposure level was chosen so that the specific energy absorption rate (SAR) would be approximately equal to the level of 4 W kg-1 which is considered by a number of organizations to be a threshold for adverse biological effects. At the end of the treatment period the mice were mated with a different group of (C3H x 101) F1 hybrid females each week for the following 8 weeks. There was no significant reduction in pregnancy rate, preimplantation survival or postimplantation survival in the exposed group compared to sham-exposed controls. At the end of the mating period a cytogenetic analysis was carried out of meiotic chromosome preparations of testicular tissue, thus sampling cells that were stem cell spermatogonia during the treatment regime. The results showed no difference in the frequency of reciprocal translocations between the sham and treated groups, or in the frequency of cells with autosome or sex chromosome univalents. Low levels of fragments and exchanges were found in both groups. It is concluded that there is no evidence in this experiment to show that chronic exposure of male mice to 2.45 GHz microwave radiation induces a mutagenic response in male germ cells. This conclusion is in agreement with the observations of Berman et al. (1980), who reported a lack of male germ cell mutagenesis after repetitive or chronic exposure of rats to 2.45 GHz.

Evaluation of lannate 20, a carbamate pesticide in the germ cells of male mice

Lannate 20, a carbamate pesticide, was evaluated for its effects on the germ cells of Swiss albino male mice, by the sperm morphology assay and meiotic chromosome preparations. The three sublethal concentrations of 20, 40, and 60 mg/kg body wt administered by the oral intubation method produced significant results (P less than 0.01) with both the above protocols in the said test system under study, thus indicating that lannate 20 is mutagenic in mice.

In VivoCytogenetic Effects of Amidopyrine + Allobarbitone on Meiotic Chromosomes of Mice

SUMMARYCytogenetic effects of an analgesic and antipyretic drug Amidopyrine + Allobarbitone were investigated in meiotic cells of Swiss albino male mice. Two sets of experiments were conducted. In one set animals received 0.65, 1.3 and 2.6 mg. doses of the drug in 0.5 ml quantities for only once (single doses series) and in other the same doses were given for thrice at 24 hr. intervals (cumulative dose series) to simulate human therapeutic conditions. Adequate controls were maintained. Meiotic chromosome preparations were made from testes after 24 hr. and at weekly intervals for five weeks. Drug exhibited a very mild clastogenic and poor c-mitotic action. It induced autosomal and sex chromosomal univalents to a significant level, the former being more. Multiple doses were found to be relatively more effective in bringing about the anomalies.

Cytogenetic Response of Meiocytes of Swiss Albino Mice to Paracetamol

SUMMARYThe in vivo cytogenetic action of the analgesic and antipyretic agent Paracetamol is assessed, for a period ranging from 24 hr. to five weeks, from meiotic cells of Swiss albino male mice administered with it at 0.625, 1.25 and 2.5 mg. doses. Two series of experiments were conducted. In one of them (single dose series) the animals received the drug only once, whereas in the other (cumulative dose series), the same concentrations were given consecutively for three days at 24 hr. intervals. Adequate controls were maintained under identical conditions. Meiotic chromosome preparations were made by the standard air drying Giemsa method after sacrificing the mice 24 hr. following drug administration and at weekly intervals. Quantitative data were subjected to statistical analysis. The compound is nonclastogenic. It induced univalents of auto- and sex chromosomal types, the latter being more frequent than autosomal ones. Polyploid cells were also observed at almost all periods. Cumulative dose administrat...

Cytogenetics of the chinese giant salamander, Andrias davidianus (Blanchard): the evolutionary significance of cryptobranchoid karyotypes

Cytogenetic aspects of the cryptobranchid salamander Andrias davidianus of western China have been studied, including chromosome number and morphology, C-band patterns, meiosis, and the chromosomal localization of ribosomal 5S RNA genes. Our data regarding chromosome number (2n=60) and general chromosome morphology largely confirm the results of Morescalchi et al. (1977). The karyotype consists of 16 pairs of “macrochromosomes” that decrease gradually in relative length to 14 pairs of “microchromosomes”. Telocentric chromosomes are a conspicuous feature of the karyotype, representing more than half the genome. Differential staining reveals that all of the chromosomes, except four pairs of microchromosomes, have C-band heterochromatin in their centromeric regions, the amount varying irrespective of chromosome size. Faint bands of interstitial and telomeric C-band heterochromatin are found in mitotic chromosomes but are not seen in meiotic preparations. In C-banded mitotic preparations from a female, one of the smallest macrochromosome pairs is heteromorphic in respect to C-band heterochromatin and centromere position. In situ hybridization of an iodinated 5S RNA probe to meiotic chromosome preparations reveals that this repeated gene is clustered near the telomeric region of chromosome 7, a medium size telocentric, a location corresponding to a band of heterochromatin. Studies of spermatocytes indicate that the process of meiosis in A. davidianus closely resembles that of more advanced salamanders, and that the microchromosomes are meiotically stable. The significance of microchromosomes and chromosome morphology in the reorganization of salamander genomes during evolution is discussed on the basis of cytogenetic data available for A. davidianus and various other primitive and advanced salamanders.

A technique for cytological preparations following enzymatic digestion of pollen mother cell walls.

A technique is presented for making meiotic chromosome preparations with enzymatic digestion of pollen mother cell walls with cellulase. This technique has given excellent chromosome definition from early pachytene stages on. Such chromosome preparations were found useful for in situ nucleic acid hybridisation studies on plant chromosomes.

Sterility and semisterility in male progeny of male mice treated with the chemical mutagen TEPA

Administration of a single subtoxic dose of TEPA [Tris(1-aziridinyl)-phosphine oxide] to 30 adult male mice induced sterility and semisterility in male F 1 and F 2 progeny of 7 treated males, during weeks 1, 2, and 3 after treatment. Examination of meiotic chromosome preparations from 4 F 2 males with reduced fertility sired by one of the semisterile F 1 males confirmed the presence of translocations.

Human meiosis: the occurrence of interlocked bivalents in a normal male.

SummaryThe mitotic and meiotic chromosomes of a healthy 65‐year‐old man, ascertained from a survey of surgically removed testes, were investigated. A method for preparation of meiotic chromosomes from testicular material is described.A pair of interlocked bivalents was observed in 9 of the 54 diakinesislearly metaphase I cells examined. This was considered an abnormally high frequency to be explained as the result of a chance event. It is suggested that the chromosomes concerned are derived from those numbered 1–3 and 13–18 in the Denver system.

Meiosis in the Human Male

An improved technique for the preparation of meiotic chromosomes from human testicular biopsy specimens for microscopic examination is described. The results are exemplified by a description of the observations of the chromosomes at various stages of the first and second meiotic divisions in a normal male.