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RNAi-based targeted gene knockdown in the model oleaginous microalgae Nannochloropsis oceanica
Wei, Li1,2; Xin, Yi1,2; Wang, Qintao1,2,3; Yang, Juan3,4; Hu, Hanhua4; Xu, Jian1,2
2017-03-01
Source PublicationPLANT JOURNAL
ISSN0960-7412
Volume89Issue:6Pages:1236-1250
AbstractMicroalgae are promising feedstock for renewable fuels such as biodiesel, yet development of industrial oleaginous strains has been hindered by the paucity and inefficiency of reverse genetics tools. Here we established an efficient RNAi-based targeted gene-knockdown method for Nannochloropsis spp., which are emerging model organisms for industrial microalgal oil production. The method achieved a 40-80% success rate in Nannochloropsis oceanica strain IMET1. When transcript level of one carbonic anhydrase (CA) was inhibited by 6283% via RNAi, mutant cells exhibited photosynthetic oxygen evolution (POE) rates that were 68-100% higher than wild-type (WT) at pH 6.0, equivalent to WT at pH 8.2, yet 39-45% lower than WT at pH 9.0. Moreover, the mutant POE rates were negatively correlated with the increase of culture pH, an exact opposite of WT. Thus, a dynamic carbon concentration mechanism (CCM) that is highly sensitive to pH homeostasis was revealed, where the CA inhibition likely partially abrogated the mechanism that normally deactivates CCM under a high level of dissolved CO2. Extension of the method to another sequenced N. oceanica strain of CCMP 1779 demonstrated comparable performance. Finally, McrBC-PCR followed by bisulfite sequencing revealed that the gene knockdown is mediated by the CG, CHG and CHH types of DNA methylation at the coding region of the targeted gene. The efficiency, robustness and general applicability of this reverse genetics approach suggested the possibility of large-scale RNAi-based gene function screening in industrial microalgae.
SubtypeArticle
KeywordRnai Oleaginous Microalgae Nannochloropsis Carbonic Anhydrase Bicarbonate Transporter Technical Advance
DOI10.1111/tpj.13411
WOS HeadingsScience & Technology ; Life Sciences & Biomedicine
Indexed BySCI
Funding OrganizationNational Natural Science Foundation of China(31425002 ; National Natural Science Foundation of China(31425002 ; Ministry of Science and Technology of China(2012CB721101 ; Ministry of Science and Technology of China(2012CB721101 ; Biological Carbon Sequestration Program from Chinese Academy of Sciences(KSZD-EW-Z-017 ; Biological Carbon Sequestration Program from Chinese Academy of Sciences(KSZD-EW-Z-017 ; Young Investigator Program from Natural Science Foundation of Shandong Province(ZR2015CQ003) ; Young Investigator Program from Natural Science Foundation of Shandong Province(ZR2015CQ003) ; 41576144) ; 41576144) ; 2012AA02A707) ; 2012AA02A707) ; ZDRW-ZS2016-3) ; ZDRW-ZS2016-3) ; National Natural Science Foundation of China(31425002 ; National Natural Science Foundation of China(31425002 ; Ministry of Science and Technology of China(2012CB721101 ; Ministry of Science and Technology of China(2012CB721101 ; Biological Carbon Sequestration Program from Chinese Academy of Sciences(KSZD-EW-Z-017 ; Biological Carbon Sequestration Program from Chinese Academy of Sciences(KSZD-EW-Z-017 ; Young Investigator Program from Natural Science Foundation of Shandong Province(ZR2015CQ003) ; Young Investigator Program from Natural Science Foundation of Shandong Province(ZR2015CQ003) ; 41576144) ; 41576144) ; 2012AA02A707) ; 2012AA02A707) ; ZDRW-ZS2016-3) ; ZDRW-ZS2016-3)
Language英语
WOS Research AreaPlant Sciences
WOS SubjectPlant Sciences
WOS KeywordDIATOM PHAEODACTYLUM-TRICORNUTUM ; CYANIDIOSCHYZON-MEROLAE 10D ; GENOME REVEALS ; HOMOLOGOUS RECOMBINATION ; HIGH-THROUGHPUT ; CHLAMYDOMONAS-REINHARDTII ; TRANSCRIPTOMIC ANALYSIS ; NUCLEAR TRANSFORMATION ; METHYLATION ANALYSIS ; FUNCTIONAL GENOMICS
WOS IDWOS:000395811000013
Funding OrganizationNational Natural Science Foundation of China(31425002 ; National Natural Science Foundation of China(31425002 ; Ministry of Science and Technology of China(2012CB721101 ; Ministry of Science and Technology of China(2012CB721101 ; Biological Carbon Sequestration Program from Chinese Academy of Sciences(KSZD-EW-Z-017 ; Biological Carbon Sequestration Program from Chinese Academy of Sciences(KSZD-EW-Z-017 ; Young Investigator Program from Natural Science Foundation of Shandong Province(ZR2015CQ003) ; Young Investigator Program from Natural Science Foundation of Shandong Province(ZR2015CQ003) ; 41576144) ; 41576144) ; 2012AA02A707) ; 2012AA02A707) ; ZDRW-ZS2016-3) ; ZDRW-ZS2016-3) ; National Natural Science Foundation of China(31425002 ; National Natural Science Foundation of China(31425002 ; Ministry of Science and Technology of China(2012CB721101 ; Ministry of Science and Technology of China(2012CB721101 ; Biological Carbon Sequestration Program from Chinese Academy of Sciences(KSZD-EW-Z-017 ; Biological Carbon Sequestration Program from Chinese Academy of Sciences(KSZD-EW-Z-017 ; Young Investigator Program from Natural Science Foundation of Shandong Province(ZR2015CQ003) ; Young Investigator Program from Natural Science Foundation of Shandong Province(ZR2015CQ003) ; 41576144) ; 41576144) ; 2012AA02A707) ; 2012AA02A707) ; ZDRW-ZS2016-3) ; ZDRW-ZS2016-3)
Citation statistics
Document Type期刊论文
Identifierhttp://ir.ihb.ac.cn/handle/342005/28957
Collection藻类生物学及应用研究中心
Affiliation1.Chinese Acad Sci, Qingdao Inst BioEnergy & Bioproc Technol, CAS Key Lab Biofuels, Single Cell Ctr, Qingdao 266101, Shandong, Peoples R China
2.Chinese Acad Sci, Qingdao Inst BioEnergy & Bioproc Technol, Shandong Key Lab Energy Genet, Qingdao 266101, Shandong, Peoples R China
3.Univ Chinese Acad Sci, Beijing 100049, Peoples R China
4.Chinese Acad Sci, Inst Hydrobiol, Key Lab Algal Biol, Wuhan 430072, Hubei, Peoples R China
Recommended Citation
GB/T 7714
Wei, Li,Xin, Yi,Wang, Qintao,et al. RNAi-based targeted gene knockdown in the model oleaginous microalgae Nannochloropsis oceanica[J]. PLANT JOURNAL,2017,89(6):1236-1250.
APA Wei, Li,Xin, Yi,Wang, Qintao,Yang, Juan,Hu, Hanhua,&Xu, Jian.(2017).RNAi-based targeted gene knockdown in the model oleaginous microalgae Nannochloropsis oceanica.PLANT JOURNAL,89(6),1236-1250.
MLA Wei, Li,et al."RNAi-based targeted gene knockdown in the model oleaginous microalgae Nannochloropsis oceanica".PLANT JOURNAL 89.6(2017):1236-1250.
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