Initiation of meiotic recombination in the yeast Saccharomyces cerevisiae occurs by localized DNA double‐strand breaks (DSBs) at several locations in the genome, corresponding to hot spots for meiotic gene conversion and crossing over. The meiotic DSBs occur in regions of chromatin that are hypersensitive to nucleases. To gain insight into the molecular mechanism involved in the formation of these DSBs, we have determined their positions at the nucleotide level at the CYS3 hot spot of gene conversion on chromosome I. We found four major new features of these DSBs: (i) sites of DSBs are multiple with varying intensities and spacing within the promoter region of the CYS3 gene; (ii) no consensus sequence can be found at these sites, indicating that the activity involved in DSB formation has little or no sequence specificity; (iii) the breaks are generated by blunt cleavages; and (iv) the 5′ ends are modified in rad50S mutant strains, where the processing of these ends is known to be prevented. We present a model for the initiation of meiotic recombination taking into account the implications of these results.

The accurate, reproducible, and timely reporting of nucleated red blood cells (NRBC) is an important function of the clinical hematology laboratory. We used 960 samples from 5 worldwide sites to evaluate a new NRBC enumeration method for the ADVIA 2120 Hematology System. The method showed excellent correlation with microscopy (r = 0.93). Sensitivity and specificity for the presence of NRBC for all samples analyzed was 77.3% and 74.6%, respectively. Almost all false negative samples were at NRBC counts 10/100 WBC. The NRBC method automatically corrects WBC counts and differential results for the presence of NRBC; the uncorrected counts are available to the user on the run screen. This method allows the enumeration of NRBC with the ADVIA 2120 Hematology System in CBC/DIFF sample mode without need for additional hardware, sample preparation, or reagents.

In Two types of meiotic recombination events have been found to occur in all eukaryotes examined, crossovers (reciprocal exchanges) and gene conversions (unidirectional transfers of information). The frequencies of both events vary within a genome (1-4). The nature of the underlying molecular processes that control the distribution of recombination events remains to be elucidated. One approach is the study of gene conversion hotspots, defined as sites or regions enhancing gene conversion in their vicinity.In fungi, where all four products of individual meiosis remain grouped in a tetrad, gene conversion events are genetically detected when a diploid strain is heterozygous at a locus (alleles A and a) leading to a tetrad containing 3A:la or 1A:3a products rather than the normal Mendelian segregation (2A:2a). In Saccharomyces cerevisiae, the frequency of meiotic gene conversion is high in the vicinity of the ARG4 gene (5, 6). The highest frequency of gene conversion (17% of total meiosis) is found for a marker located in the ARG4 promoter region around position -119 (Fig. 1). The frequencies decrease in a gradient, termed polarity, for markers located on either side of the ARG4 promoter (10). Within the ARG4 coding region, the conversion frequencies decrease from 10% (position +3) to 0.4% (position +1274). Two previous deletion analyses mapped an initiation site for meiotic gene conversion in the promoter region of ARG4 between positions -316 and -37 (6, 7). These genetics data and the physical analysis of the DNA of this chromosomal region during meiosis, which demonstrated the occurrence of a transient double-strand break (DSB) around position -200 (8, 9, 11), strongly suggest that the implicated region includes a cis-acting initiation site which stimulates recombination in its vicinity.To know whether all the cis-acting elements had been identified by deletion analysis, we tested whether the -319/ -37 sequence was sufficient to promote gene conversion in a novel chromosomal context-i.e., when displaced so that it was adjacent to new flanking sequences. We chose to invert the ARG4 gene on the chromosome. This "minimal displacement" was preferred to the random insertion of DNA fragments on different chromosomes, which is known to be strongly subject to position effects for recombination between LEU2 (12) or ARG4 (13) heteroalleles. However, a 12-kilobase (kb) ARG4-containing fragment inserted on a yeast artificial chromosome has the same recombinational properties as at its normal location on chromosome VIII (14).We report here that the necessary -316/-37 cis-acting sequence previously defined by deletion analysis at the normal location is not sufficient to promote gene conversion at ARG4 in any chromosomal context. We show that the presence of a transcriptional terminator upstream from the initiation site in these inversions is absolutely required in order to maintain both the DSB and the recombination properties at ARG4. We also show that high and low levels of gene conversion at ARG4 obtained in variou...

Background: Meiotic recombination between homologous chromosomes in the yeast Saccharo-myces cerevisiae is initiated by the formation of DNA double-strand breaks (DSBs). The mechanism of DSB formation and the factors that determine their frequency and location have yet to be elucidated. Current studies of meiotic recombination are also concerned with the question of the functional relationship between DSB formation and the other meiotic processes of homology searching, pairing and synapsis of homologues.

In prokaryotes, the RecA protein plays a pivotal role in homologous recombination, catalyzing the transfer of a single DNA strand into an homologous molecule. Structural homologs of the bacterial RecA protein, called Rad51, have been found in different eukaryotes (from yeast to man), suggesting a certain level of conservation in recombination pathways among living organisms. We have cloned the homolog of RAD51 in Caenorhabditis elegans. The CeRAD51 gene is transcribed into two alternative mRNAs and potentially codes for two proteins of 395 and 357 amino acids in length, respectively. We discuss the evolutionary implications of these findings.

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Imerslund-Gräsbeck syndrome is a rare autosomal recessive disorder, characterized by vitamin B12 deficiency due to selective malabsorption of the vitamin and usually results in megaloblastic anemia appearing in childhood. It is responsive to parenteral vitamin B12 therapy.The estimated prevalence (calculated based on Scandinavian data) is less than 6:1,000,000. However, many cases may be misdiagnosed.When there is reasonable evidence to suspect that a patient suffers from IGS, a new and straightforward approach to diagnosis is mutational analysis of the appropriate genes. We report for the first time the case of a girl of Italian ancestry with IGS genetically confirmed by the detection of a homozygous missense mutation in the AMN gene (c.208-2 A > G).

We evaluated an extended time in the microscopic review in samples in which the potential clinical information could be increased with respect to those that could be achieved with the usual laboratory methodologies. We used samples containing nucleated red blood cells in a small amount and cytopenic samples. For these purposes for each peripheral blood smear, the timing of eye-count differential was increased up to 20 min, regardless of the final number of cells which could be counted. In addition, an automated system for digital analysis of peripheral blood smears was employed and the number of cells counted was brought up to 1000 leukocytes. In both manual and automatic light microscopy extended observation, we obtained more diagnostic information in respect to those with routine or standard methods. Both automated and manual increase systems of the timing for microscopic review are useful tools to find diagnostic information that otherwise would be lost using normal and standard procedures. So, these methods should be used especially when there is a higher pre-test probability for discovery of pathological cells.