Simplified All-In-One CRISPR-Cas9 Construction for Efficient Genome Editing in Cryptococcus Species.
Simplified All-In-One CRISPR-Cas9 Construction for Efficient Genome Editing in Cryptococcus Species.
复制标题
简化的一体式 CRISPR-Cas9 构建，可在隐球菌属物种中进行高效基因组编辑
DOI：
10.3390/jof7070505
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发表时间：
2021-06-24
期刊：
影响因子：
--
通讯作者：
Zhu X
中科院分区：
文献类型：
--
作者：
Zhang P;Wang Y;Li C;Ma X;Ma L;Zhu X
Cryptococcus neoformans and Cryptococcus deneoformans are opportunistic fungal pathogens found worldwide that are utilized to reveal mechanisms of fungal pathogenesis. However, their low homologous recombination frequency has greatly encumbered genetic studies. In preliminary work, we described a ‘suicide’ CRISPR-Cas9 system for use in the efficient gene editing of C. deneoformans, but this has not yet been used in the C. neoformans strain. The procedures involved in constructing vectors are time-consuming, whether they involve restriction enzyme-based cloning of donor DNA or the introduction of a target sequence into the gRNA expression cassette via overlap PCR, as are sophisticated, thus impeding their widespread application. Here, we report the optimized and simplified construction method for all-in-one CRISPR-Cas9 vectors that can be used in C. neoformans and C. deneoformans strains respectively, named pNK003 (Genbank: MW938321) and pRH003 (Genbank: KX977486). Taking several gene manipulations as examples, we also demonstrate the accuracy and efficiency of the new simplified all-in-one CRISPR-Cas9 genome editing tools in both Serotype A and Serotype D strains, as well as their ability to eliminate Cas9 and gDNA cassettes after gene editing. We anticipate that the availability of new vectors that can simplify and streamline the technical steps for all-in-one CRISPR-Cas9 construction could accelerate genetic studies of the Cryptococcus species.
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影响因子：
46.9
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影响因子：
64.8
作者：
MANSOUR, SL;THOMAS, KR;CAPECCHI, MR
通讯作者：
CAPECCHI, MR
影响因子：
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作者：
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通讯作者：
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DOI：
10.1084/jem.184.2.377
发表时间：
1996-08-01
期刊：
The Journal of experimental medicine
影响因子：
--
作者：
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通讯作者：
Williamson PR