Biolistic Transformation of Haematococcus pluvialis With Constructs Based on the Flanking Sequences of Its Endogenous Alpha Tubulin Gene

doi:10.3389/fmicb.2019.01749

CORC > 烟台海岸带研究所 > 中国科学院烟台海岸带研究所 > 海岸带生物学与生物资源利用重点实验室 > 海岸带生物学与生物资源利用所重点实验室

	Biolistic Transformation of Haematococcus pluvialis With Constructs Based on the Flanking Sequences of Its Endogenous Alpha Tubulin Gene
	Yuan, GH ; Xu, XY ; Zhang, W ; Zhang, WL ; Cui, YL ; Qin, S ; Liu, TZ
刊名	FRONTIERS IN MICROBIOLOGY
	2019-08-02
卷号	10 页码:1749
关键词	alpha tubulin biolistic transformation flanking sequence Haematococcus pluvialis selection marker
ISSN号	1664-302X
DOI	10.3389/fmicb.2019.01749
产权排序	[Yuan, Guanhua ; Xu, Xiaoying ; Zhang, Wei ; Zhang, Wenlei ; Liu, Tianzhong] Chinese Acad Sci, Qingdao Inst Bioenergy & Bioproc Technol, Key Lab Biofuels, Qingdao, Shandong, Peoples R China ; [Yuan, Guanhua ; Zhang, Wei ; Zhang, Wenlei ; Liu, Tianzhong] Chinese Acad Sci, Qingdao Inst Bioenergy & Bioproc Technol, Shandong Prov Key Lab Energy Genet, Qingdao, Shandong, Peoples R China ; [Yuan, Guanhua] Univ Chinese Acad Sci, Beijing, Peoples R China ; [Xu, Xiaoying] Yantai Marine Econ Res Inst, Yantai, Peoples R China ; [Cui, Yulin ; Qin, Song] Chinese Acad Sci, Yantai Inst Coastal Zone Res, Key Lab Coastal Biol & Biol Resource Utilizat, Yantai, Peoples R China
文献子类	Article
英文摘要	Haematococcus pluvialis has high commercial value, yet it displays low development of genetic transformation systems. In this research, the endogenous 5' and 3' flanking sequences of the constitutive alpha tubulin (tub) gene were cloned along with its encoding region in H. pluvialis, in which some putative promoter elements and polyadenylation signals were identified, respectively. Three selection markers of tub/aadA, tub/hyr and tub/ble with three different antibiotic-resistance genes fused between the endogenous tub promoter (Ptub) and terminator (Ttub) were constructed and utilized for biolistic transformation of H. pluvialis. Stable resistant colonies with introduced aadA genes were obtained after bombardments of either H. pluvialis NIES144 or SCCAP K0084 with the tub/aadA cassette, the efficiency of which could reach up to 3 x 10(-5) per mu g DNA through an established manipulation flow. Two key details, including the utilization of culture with motile flagellates dominant and controlled incubation of them on membrane filters during bombardments, were disclosed firstly. In obtained transformants, efficient integration and transcription of the foreign tub/aadA fragments could be identified through genome PCR examination and qPCR analysis, nonetheless with random style instead of homologous crossover in the H. pluvialis genome. The presented selection marker and optimized transforming procedures in this report would strengthen the platform for genetic manipulation and modification of H. pluvialis.
WOS关键词	NUCLEAR TRANSFORMATION ; ASTAXANTHIN ; EXPRESSION ; PROMOTER ; CHLOROPHYCEAE ; PCR ; RECOMBINATION ; ELEMENTS ; GROWTH
WOS研究方向	Microbiology
语种	英语
WOS记录号	WOS:000478564700001
资助机构	2016 Sino-Thai Cooperation Project by the National Natural Science Foundation of ChinaNational Natural Science Foundation of China [51561145015] ; Shandong Provincial Key Laboratory of Energy Genetics [SDKLEG201805] ; Synthetic Biology Technology Innovation Center of Shandong Province [sdsynbio-2018-PY-08]
内容类型	期刊论文
源URL	[http://ir.yic.ac.cn/handle/133337/24885]
专题	烟台海岸带研究所_海岸带生物学与生物资源利用所重点实验室
作者单位	1.Univ Chinese Acad Sci, Beijing, Peoples R China; 2.Chinese Acad Sci, Qingdao Inst Bioenergy & Bioproc Technol, Key Lab Biofuels, Qingdao, Shandong, Peoples R China; 3.Chinese Acad Sci, Qingdao Inst Bioenergy & Bioproc Technol, Shandong Prov Key Lab Energy Genet, Qingdao, Shandong, Peoples R China; 4.Yantai Marine Econ Res Inst, Yantai, Peoples R China; 5.Chinese Acad Sci, Yantai Inst Coastal Zone Res, Key Lab Coastal Biol & Biol Resource Utilizat, Yantai, Peoples R China
推荐引用方式 GB/T 7714	Yuan, GH,Xu, XY,Zhang, W,et al. Biolistic Transformation of Haematococcus pluvialis With Constructs Based on the Flanking Sequences of Its Endogenous Alpha Tubulin Gene[J]. FRONTIERS IN MICROBIOLOGY,2019,10:1749.
APA	Yuan, GH.,Xu, XY.,Zhang, W.,Zhang, WL.,Cui, YL.,...&Liu, TZ.(2019).Biolistic Transformation of Haematococcus pluvialis With Constructs Based on the Flanking Sequences of Its Endogenous Alpha Tubulin Gene.FRONTIERS IN MICROBIOLOGY,10,1749.
MLA	Yuan, GH,et al."Biolistic Transformation of Haematococcus pluvialis With Constructs Based on the Flanking Sequences of Its Endogenous Alpha Tubulin Gene".FRONTIERS IN MICROBIOLOGY 10(2019):1749.