Genetic characteristics and ploidy trigger the high inducibility of double haploid (DH) inducer in Brassica napus

Background Our recently reported doubled haploid (DH) induction lines e.g., Y3380 and Y3560 are allo-octoploid (AAAACCCC, 2n = 8× ≈ 76), which can induce the maternal parent to produce DH individuals. Whether this induction process is related to the production of aneuploid gametes form male parent and genetic characteristics of the male parent has not been reported yet. Results Somatic chromosome counts of DH inducer parents, female wax-less parent (W1A) and their F1 hybrid individuals revealed the reliability of flow cytometry analysis. Y3560 has normal chromosome behavior in metaphase I and anaphase I, but chromosome division was not synchronized in the tetrad period. Individual phenotypic identification and flow cytometric fluorescence measurement of F1 individual and parents revealed that DH individuals can be distinguished on the basis of waxiness trait. The results of phenotypic identification and flow cytometry can identify the homozygotes or heterozygotes of F1 generation individuals. The data of SNP genotyping coupled with phenotypic waxiness trait revealed that the genetic distance between W1A and F1 homozygotes were smaller as compared to their heterozygotes. It was found that compared with allo-octoploids, aneuploidy from allo-octoploid segregation did not significantly increase the DH induction rate, but reduced male infiltration rate and heterozygous site rate of induced F1 generation. The ploidy, SNP genotyping and flow cytometry results cumulatively shows that DH induction is attributed to the key genes regulation from the parents of Y3560 and Y3380, which significantly increase the induction efficiency as compared to ploidy. Conclusion Based on our findings, we hypothesize that genetic characteristics and aneuploidy play an important role in the induction of DH individuals in Brassca napus, and the induction process has been explored. It provides an important insight for us to locate and clone the genes that regulate the inducibility in the later stage.

Loss of CEP70 function affects acrosome biogenesis and flagella formation during spermiogenesis

The spermatogenesis process is complex and delicate, and any error in a step may cause spermatogenesis arrest and even male infertility. According to our previous transcriptomic data, CEP70 is highly expressed throughout various stages of human spermatogenesis, especially during the meiosis and deformation stages. CEP70 is present in sperm tails and that it exists in centrosomes as revealed by human centrosome proteomics. However, the specific mechanism of this protein in spermatogenesis is still unknown. In this study, we found a heterozygous site of the same mutation on CEP70 through mutation screening of patients with clinical azoospermia. To further verify, we deleted CEP70 in mice and found that it caused abnormal spermatogenesis, leading to male sterility. We found that the knockout of CEP70 did not affect the prophase of meiosis I, but led to male germ-cell apoptosis and abnormal spermiogenesis. By transmission electron microscopy (TEM) and scanning electron microscopy (SEM) analysis, we found that the deletion of CEP70 resulted in the abnormal formation of flagella and acrosomes during spermiogenesis. Tandem mass tag (TMT)-labeled quantitative proteomic analysis revealed that the absence of CEP70 led to a significant decrease in the proteins associated with the formation of the flagella, head, and acrosome of sperm, and the microtubule cytoskeleton. Taken together, our results show that CEP70 is essential for acrosome biogenesis and flagella formation during spermiogenesis.

Variable Spontaneous Mutation and Loss of Heterozygosity among Heterozygous Genomes in Yeast

Mutation and recombination are the primary sources of genetic variation. To better understand the evolution of genetic variation, it is crucial to comprehensively investigate the processes involving mutation accumulation and recombination. In this study, we performed mutation accumulation experiments on four heterozygous diploid yeast species in the Saccharomycodaceae family to determine spontaneous mutation rates, mutation spectra, and losses of heterozygosity (LOH). We observed substantial variation in mutation rates and mutation spectra. We also observed high LOH rates (1.65–11.07×10−6 events per heterozygous site per cell division). Biases in spontaneous mutation and LOH together with selection ultimately shape the variable genome-wide nucleotide landscape in yeast species.

Heterozygosity increases microsatellite mutation rate

Whole genome sequencing of families of Arabidopsis has recently lent strong support to the heterozygote instability (HI) hypothesis that heterozygosity locally increases mutation rate. However, there is an important theoretical difference between the impact on base substitutions, where mutation rate increases in regions surrounding a heterozygous site, and the impact of HI on sequences such as microsatellites, where mutations are likely to occur at the heterozygous site itself. At microsatellite loci, HI should create a positive feedback loop, with heterozygosity and mutation rate mutually increasing each other. Direct support for HI acting on microsatellites is limited and contradictory. I therefore analysed AC microsatellites in 1163 genome sequences from the 1000 genomes project. I used the presence of rare alleles, which are likely to be very recent in origin, as a surrogate measure of mutation rate. I show that rare alleles are more likely to occur at locus-population combinations with higher heterozygosity even when all populations carry exactly the same number of alleles.

A general method for identifying correct solutions in the quantitative analysis of gene conversion data

Past attempts to obtain values for meiotic parameters relating to hybrid DNA formation and the correction of mismatched bases in hybrid DNA have not given unique solutions unless various simplifying assumptions were made. A method is given for identifying correct sets of solutions after calculating the frequency of hybrid DNA formation at a heterozygous site and using the fact that closely linked sites within a locus have very similar hybrid DNA formation frequencies. The method is illustrated with simulated data and Sordaria fimicola data; it can also show up incorrect assumptions in analysis. A method is suggested for assessing the importance of double-strand gaps in producing conversions.Key words: recombination, gene conversion, quantitative analysis.

The interpretation of gene conversion data from ordered eight-spored asci

SUMMARYCurrently favoured models postulate that gene conversion is due to the correction of mis-matches in heteroduplex DNA. If heteroduplex is formed reciprocally on both chromatids participating in recombination, the mis-matches due to a heterozygous site will be different on the two chromatids, and there will be four correction probabilities to be taken into account. It is shown that, given the frequencies of the five different kinds of aberrant ascus ratios, it is possible to calculate four alternative sets of values for the four correction probabilities and the total number of asci in which heteroduplex is formed. These four solutions reduce in effect to two when there are no other markers distinguishing the two chromatids. With the aid of flanking markers and the assumption that heteroduplex formation is chemically polarized, it is possible, in principle, to choose one best solution.The method has been applied to the five one-point crosses in Sordaria fimicola from which most data are available. The data from four different mutants crossed to wild type are compatible with a restricted model in which the correction frequencies, from mutant to wild and from wild to mutant, are the same on both chromatids. In the case of the fifth mutant the data are not consistent with this restricted model, and indicate different correction frequencies in the two chromatids.

A mechanism for gene conversion in fungi

A mechanism for gene conversion is proposed which overcomes many of the difficulties that any copy choice model encounters. It is suggested that along with general genetic pairing of homologous genomes at meiosis, effective pairing over short regions of the genetic material occurs at the molecular level by the separation of the strands of the DNA double helices, followed by the annealing of strands from two homologous chromatids. If the annealed region happens to span a heterozygous site, mispairing of bases will occur. Such a situation may be analogous to that in DNA which is damaged by mutagens; the same or similar repair mechanisms may operate, and these, by adjusting the base sequences in order to restore normal base pairing, would bring about gene conversion in the absence of any genetic replication. The model indicates how precise breakage and rejoining of chromatids could occur in the vicinity of the conversion, so that conversion would frequently be accompanied by the recombination of outside markers. The model also proposes that the distance between two mutant sites on a fine structure map depends not so much on the frequency of a recombinational event occurring between them, but rather on the degree of inhibition of the processes of genetic pairing by the mutants themselves.The model will explain almost all the data in a formal way, and it has the advantage over copy choice mechanisms for gene conversion in (1) being compatible with semi-conservative replication of DNA, (2) not invoking DNA synthesis during or after genetic pairing, (3) providing a molecular mechanism for close specific pairing, (4) making it unnecessary to postulate sister strand exchange or a process akin to this, (5) suggesting why rates of gene conversion in opposite directions are sometimes unequal and (6) providing an explanation of the clustering of mutant sites, a basis for map expansion and for the apparently capricious departure of fine structure maps from additivity. Although the model proposed is a general rather than a specific one, it suggests that the process of conversion and intragenic recombination is more complex than is usually believed, since it depends on several interacting factors. Nevertheless, it is hoped that the introduction of a model with this complexity will help to stimulate specific experiments, and that these will provide definitive information which would never be obtained if simpler models of conversion and intragenic recombination were believed to explain the genetic data sufficiently well.