Chemical transformation of the multibudding yeast, Aureobasidium pullulans.

IF 7.4 1区 生物学 Q1 CELL BIOLOGY Journal of Cell Biology Pub Date : 2024-10-07 Epub Date: 2024-06-27 DOI:10.1083/jcb.202402114
Alison C E Wirshing, Claudia A Petrucco, Daniel J Lew
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Abstract

Aureobasidium pullulans is a ubiquitous polymorphic black yeast with industrial and agricultural applications. It has recently gained attention amongst cell biologists for its unconventional mode of proliferation in which multinucleate yeast cells make multiple buds within a single cell cycle. Here, we combine a chemical transformation method with genome-targeted homologous recombination to yield ∼60 transformants/μg of DNA in just 3 days. This protocol is simple, inexpensive, and requires no specialized equipment. We also describe vectors with codon-optimized green and red fluorescent proteins for A. pullulans and use these tools to explore novel cell biology. Quantitative imaging of a strain expressing cytosolic and nuclear markers showed that although the nuclear number varies considerably among cells of similar volume, total nuclear volume scales with cell volume over an impressive 70-fold size range. The protocols and tools described here expand the toolkit for A. pullulans biologists and will help researchers address the many other puzzles posed by this polyextremotolerant and morphologically plastic organism.

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多芽酵母 Aureobasidium pullulans 的化学转化。
拉氏酵母菌(Aureobasidium pullulans)是一种普遍存在的多态黑色酵母菌,在工业和农业领域都有应用。最近,它因其非传统的增殖模式(多核酵母细胞在一个细胞周期内产生多个芽)而受到细胞生物学家的关注。在这里,我们将化学转化方法与基因组靶向同源重组相结合,仅用 3 天就能获得 ∼60 个转化子/μg DNA。这种方法简单、廉价,而且不需要专门的设备。我们还描述了带有密码子优化的绿色和红色荧光蛋白的拉氏杆菌载体,并利用这些工具探索新的细胞生物学。对一株表达细胞膜和细胞核标记物的菌株进行定量成像显示,虽然细胞核数量在体积相似的细胞中变化很大,但细胞核的总体积与细胞体积成比例,大小范围达到令人印象深刻的 70 倍。本文描述的方案和工具扩展了 A. pullulans 生物学家的工具包,将帮助研究人员解决这种多极端耐受性和形态可塑性生物带来的许多其他难题。
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来源期刊
Journal of Cell Biology
Journal of Cell Biology 生物-细胞生物学
CiteScore
12.60
自引率
2.60%
发文量
213
审稿时长
1 months
期刊介绍: The Journal of Cell Biology (JCB) is a comprehensive journal dedicated to publishing original discoveries across all realms of cell biology. We invite papers presenting novel cellular or molecular advancements in various domains of basic cell biology, along with applied cell biology research in diverse systems such as immunology, neurobiology, metabolism, virology, developmental biology, and plant biology. We enthusiastically welcome submissions showcasing significant findings of interest to cell biologists, irrespective of the experimental approach.
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