IHB OpenIR  > 藻类生物学及应用研究中心  > 期刊论文
Building a multipurpose insertional mutant library for forward and reverse genetics in Chlamydomonas
Cheng, Xi1; Liu, Gai1; Ke, Wenting1; Zhao, Lijuan1,2; Lv, Bo1,2; Ma, Xiaocui1,2; Xu, Nannan1,2; Xia, Xiaoling1; Deng, Xuan1; Zheng, Chunlei3; Huang, Kaiyao1
2017-05-15
Source PublicationPLANT METHODS
ISSN1746-4811
Volume13Issue:1Pages:16
Abstract

Background: The unicellular green alga, Chlamydomonas reinhardtii, is a classic model for studying flagella and biofuel. However, precise gene editing, such as Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) and CRISPR-associated protein (Cas9) system, is not widely used in this organism. Screening of random insertional mutant libraries by polymerase chain reaction provides an alternate strategy to obtain null mutants of individual gene. But building, screening, and maintaining such a library was time-consuming and expensive. Results: By selecting a suitable parental strain, keeping individual mutants using the agar plate, and designing an insertion cassette-specific primer for library screening, we successfully generated and maintained similar to 150,000 insertional mutants of Chlamydomonas, which was used for both reverse and forward genetics analysis. We obtained 26 individual mutants corresponding to 20 genes and identified 967 motility-defect mutants including 10 mutants with defective accumulation of intraflagellar transport complex at the basal body. We also obtained 929 mutants defective in oil droplet assembly after nitrogen deprivation. Furthermore, a new insertion cassette with splicing donor sequences at both ends was also constructed, which increased the efficiency of gene interruption. Conclusion: In summary, this library provides a multifunctional platform both for obtaining mutants of interested genes and for screening of mutants with specific phenotype.

KeywordChlamydomonas Insertional mutants Mutant library Flagella Intraflagellar transport Oil droplet
DOI10.1186/s13007-017-0183-5
Funding OrganizationNational Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China
Indexed BySCI
Language英语
Funding ProjectNational Natural Science Foundation of China[31371354] ; National Natural Science Foundation of China[31400654]
WOS Research AreaBiochemistry & Molecular Biology ; Plant Sciences
WOS SubjectBiochemical Research Methods ; Plant Sciences
WOS IDWOS:000401181400001
WOS KeywordNUCLEAR TRANSFORMATION ; SELECTABLE MARKER ; GREEN-ALGA ; REINHARDTII ; EXPRESSION ; MODEL ; RESISTANCE ; PROTEINS ; DNA ; COMPLEMENTATION
PublisherBIOMED CENTRAL LTD
Funding OrganizationNational Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China
Citation statistics
Cited Times:11[WOS]   [WOS Record]     [Related Records in WOS]
Document Type期刊论文
Identifierhttp://ir.ihb.ac.cn/handle/342005/32535
Collection藻类生物学及应用研究中心_期刊论文
Corresponding AuthorHuang, Kaiyao
Affiliation1.Chinese Acad Sci, Inst Hydrobiol, Key Lab Algal Biol, Wuhan 430072, Peoples R China
2.Univ Chinese Acad Sci, Beijing 100039, Peoples R China
3.Hubei Univ, Coll Life Sci, Wuhan 430062, Peoples R China
Recommended Citation
GB/T 7714
Cheng, Xi,Liu, Gai,Ke, Wenting,et al. Building a multipurpose insertional mutant library for forward and reverse genetics in Chlamydomonas[J]. PLANT METHODS,2017,13(1):16.
APA Cheng, Xi.,Liu, Gai.,Ke, Wenting.,Zhao, Lijuan.,Lv, Bo.,...&Huang, Kaiyao.(2017).Building a multipurpose insertional mutant library for forward and reverse genetics in Chlamydomonas.PLANT METHODS,13(1),16.
MLA Cheng, Xi,et al."Building a multipurpose insertional mutant library for forward and reverse genetics in Chlamydomonas".PLANT METHODS 13.1(2017):16.
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