bioRxiv Science⌕ Search
Home/Archive/bioRxiv

bioRxiv · 10.1101/704627

Tetrad analysis without tetrad dissection: Meiotic recombination and genomic diversity in the yeast Komagataella phaffii (Pichia pastoris)

Braun-Galleani, S.·Dias, J. A.·Coughlan, A. Y.·Ryan, A. P.·Byrne, K. P.·Wolfe, K. H.
View original source ↗Export metadata ↓

Abstract

Komagataella phaffii is a yeast widely used in the pharmaceutical and biotechnology industries, and is one of the two species that were formerly called Pichia pastoris. However, almost all laboratory work on K. phaffii has been done on strains derived from a single natural isolate, CBS7435. There is little information about the genetic properties of K. phaffii or its sequence diversity. Genetic analysis is difficult because, although K. phaffii makes asci with four spores, the spores are small and tend to clump together, making the asci hard to dissect. Here, we sequenced the genomes of all the known isolates of this species, and find that K. phaffii has only been isolated from nature four times. We analyzed the meiotic recombination landscape in a cross between auxotrophically marked strains derived from two isolates that differ at 44,000 single nucleotide polymorphism sites. We conducted tetrad analysis by making use of the property that haploids of this species do not mate in rich media, which enabled us to isolate and sequence the four types of haploid cell that are present in the colony that forms when a tetratype ascus germinates. We found that approximately 25 crossovers occur per meiosis, which is 3.5 times fewer than in Saccharomyces cerevisiae. Recombination is suppressed, and genetic diversity among natural isolates is low, in a region around centromeres that is much larger than the centromeres themselves. Our method of tetrad analysis without tetrad dissection will be applicable to other species whose spores do not mate spontaneously after germination.\n\nAuthor summaryTo better understand the basic genetics of the budding yeast Komagataella phaffii, which has many applications in biotechnology, we investigated its genetic diversity and its meiotic recombination landscape. We made a genetic cross between strains derived from two natural isolates, and developed a method for characterizing the genomes of the four spores resulting from meiosis, which were previously impossible to isolate. We found that K. phaffii has a lower recombination rate than Saccharomyces cerevisiae. It shows a large zone of suppressed recombination around its centromeres, which may be due to the structural differences between centromeres in K. phaffii and S. cerevisiae.

Explore related subjects

geneticsPreprint

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Braun-Galleani, S., Dias, J. A., Coughlan, A. Y., Ryan, A. P., Byrne, K. P., Wolfe, K. H.. 2019-07-16. Tetrad analysis without tetrad dissection: Meiotic recombination and genomic diversity in the yeast Komagataella phaffii (Pichia pastoris). https://doi.org/10.1101/704627

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

The histone demethylase Kdm5 and the ARGONAUTE proteins Piwi and Aubergine regulate female abdominal pigmentation in Drosophila melanogaster

Insect pigmentation is an ecologically critical trait influencing many physiological processes. In Drosophila melanogaster, abdominal pigmentation is sexually dimorphic: males have fully pigmented posterior segments, while females exhibit a posterior melanin stripe. Pigmentation relies on the expression of pigmentation genes that encode enzymes involved in pigment synthesis. These genes are tightly regulated during pupal and young adult stages. To expand the gene regulatory network of pigmentation genes, we conducted an RNAi screen using the yellow-Gal4 driver, expressed during the pupal stage in abdominal epidermis. One of the candidates from this screen, Kdm5, encodes a histone demethylase erasing the H3K4me3 histone mark catalyzed by the histone methyl-transferase Trithorax (Trx). We show that Kdm5 down-regulation reduces abdominal pigmentation, mimicking trx down-regulation. Kdm5 activates melanin production through regulation of the pigmentation gene tan. Transcriptomic analyses reveal that Kdm5 and Trx share many targets in pupal abdominal epidermis, including piRNA pathway components such as piwi and aubergine. These piRNA components, originally associated with transposon silencing in the germline, also function in some somatic tissues such as the nervous system, the fat body or the gut. We demonstrate that Piwi and Aubergine participate in female abdominal pigmentation establishment, without evident piRNA production. We also show that Kdm5 and Piwi act not only in pupal abdominal epidermis but also in pupal fat body. This study therefore expands the regulatory network of pigmentation genes. It identifies a new somatic function for Kdm5 and Piwi and reveals a role for pupal fat body in female abdominal pigmentation regulation.

genetics↗
bioRxiv2026

Genetic diversity within and between polyploid sugarcane (Saccharum spp.) families obtained via caryopsis using microsatellite markers and multicategory model

Genetic diversity analyses are essential for sugarcane (Saccharum spp.) breeding programs. Crossbreeding, based on genetic distances between parental plants, is a tool used to increase genetic variability and enhance plant selection; however, quantifying variation in highly polyploid species remains a challenge. The present study aimed to evaluate the diversity within and between 12 families of sugarcane derived from caryopses, analyzing 120 individual seedlings arranged in an augmented block design. Genotyping was performed using primers for 16 microsatellite loci, five simple sequence repeat (SSR) loci, and 11 expressed sequence tag-SSR (EST-SSR) loci. To accurately account for polyploidy, similarity calculations were performed using Bruvos distances among individuals and RST distances among the families. Analysis of molecular variance (AMOVA) indicated that most of the genetic variability was within families (72%), with only 28% found between them. This high level of intra-family variation demonstrates that a significant reservoir of genetic diversity remains available within the crosses. The highest genetic similarity was observed between the families RB986952 x RB986960 and RB036122 x RB03611, whereas the lowest genetic similarity was observed between the families RB97319 x RB966928 and RB106802 x RB855036. Although the evaluated families shared high genetic similarity, the pronounced genetic variation within them demonstrates a robust recombination potential, indicating that the genetic basis of sugarcane can be better explored using the high variability that already exists in the selection of desirable morpho-agronomic characteristics within the families. Furthermore, this study highlights the importance of using appropriate distances for diversity studies with codominant markers, such as microsatellites, in polyploid species.

genetics↗
bioRxiv2026

Optimizing DNA extraction from environmentally degraded bone samples for molecular identification of cetacean species

Molecular identification of cetacean bone remains can be limited by DNA degradation and the presence of PCR inhibitors. Here, we present an optimized DNA extraction protocol based on a total demineralization method for environmentally exposed cetacean bones. The protocol uses 100 mg of bone powder, 24 h digestion with EDTA, N-lauroylsarcosine, and proteinase K, followed by a modified silica-column purification. Nine environmentally degraded bone samples representing eight individuals were processed. DNA concentrations ranged from 7.3 to 57.1 ng/uL (mean SD = 25.91- 13.91 ng/uL). The mitochondrial cytochrome b gene was successfully amplified from all samples using conventional PCR, and five samples (55.6%) yielded sequences suitable for downstream analysis. BLASTn identified Balaenoptera physalus as the closest database match for all recovered sequences, and phylogenetic analysis further supported their association with B. physalus reference sequences. These results demonstrate that the proposed protocol provides a practical approach for recovering amplifiable and molecularly informative mitochondrial DNA from environmentally degraded cetacean bone material, facilitating molecular identification from challenging skeletal remains.

genetics↗