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A Novel Chimeric Mitochondrial Gene Confers Cytoplasmic Effects on Seed Oil Content in Polyploid Rapeseed (Brassica napus)

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Molecular Plant 2019年第4期12卷 582-596页

作者： Jun Liu Wanjun Hao Jing Liu shihang fan Wei Zhao Linbing Deng Xinfa Wang Zhiyong Hu Wei Hua Hanzhong Wang Key Laboratory of Biology and Genetic Improvement of Oil Crops Ministry of Agriculture and Rural Affairs Oil Crops Research Institute of the Chinese Academy of Agricultural Sciences Wuhan 430062 China Key Laboratory of Tea Biology and Resources Utilization Ministry of Agriculture and Rural Affairs Tea Research Institute of the Chinese Academy of Agricultural Sciences Hangzhou 310008 China

Cytoplasmic effects (CEs) have been discovered to influence a diverse array of agronomic traits in crops, and understanding the underlying mechanisms can help accelerate breeding programs. Seed oil content (SOC) is of great agricultural, nutritional, and economic importance. However, the genetic basis of CEs on SOC (CE-SOC) remains enigmatic. In this study, we use an optimized approach to sequence the cytoplasmic (plastid and mitochondrial) genomes of allotetraploid oilseed rape (Brassica napus) cultivars, 51218 and 56366, that bear contrasting CE-SOC. By combining comparative genomics and genome-wide transcriptome analysis, we identify mitochondria-encoded orf188 as a potential CE-SOC determinant gene. Functional analyses in the model system Arabidopsis thaliana and rapeseed demonstrated that orf188 governs CE-SOC and could significantly increase SOC, strikingly, through promoting the yield of ATP. Consistent with this finding, transcriptional profiling with microarray and RNA sequencing revealed that orf188 affects transcriptional reprogramming of mitochondrial energy metabolism to facilitate ATP production. Intriguingly, orf188 is a previously uncharacterized chimeric gene, and the presence of this genetic novelty endows rapeseed with positive CE-SOC. Our results shed light on the molecular basis of CEs on a key quantitative trait in polyploid crops and enrich the theory of maternal control of oil content, providing new scientific guidance for breeding high-oil rapeseed germplasms.

关键词： cytoplasmic effects seed oil content energy metabolism crop breeding Brassica napus

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Synthesis and characteristic of a novel green fluorescent protein eYGFPuv

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Oil Crop Science 2019年第2期4卷 65-74页

作者： shihang fan Linbin Deng Hongli Yang Liang Zhang Xingchao Sun Jinglin Liu Jing Liu Key Laboratory for Biological Sciences of Oil Crops Ministry of Agriculture Oil Crops Research Institute Chinese Academy of Agriculture Sciences

Recently, a novel green fluorescent protein eYGFPuv has been identified in the marine organism Chiridius poppei which displays high fluorescence intensity and can be visible by eyes in dark. Although strong green fluorescence was achieved in transgenic petunia, 3 expression cassettes (about 8 kb) complicate its application. In this study, to confirm whether 1 expression cassette could be used as a transgenic marker in prokaryotes and eukaryotes, eYGFPuv was cloned into prokaryotic expression vector pET28α-eYGFPuv- His and plant binary expression vector 35S::eYGFPuv. Compared to EGFP, eYGFPuv protein exhibited stronger dazzling green fluorescence in E. coli under excited light at 365 nm and maintains steadily over a long period of time without degradation. When transiently expressed in tobacco leaves, eYGFPuv protein displayed strong green fluorescence. Moreover, the fluorescence of eYGFPuv protein also could be directly observed in living plant, and thus can be used easily as a marker to screen transformed lines in transgenic research. Overall, compared to previous studies on eYGFPuv tandem repeats, our data confirmed that single eYGFPuv sequence still possesses high fluorescence intensity and quenching resistance. Furthermore, because of small size of expression cassette,it is suitable for efficient transformation in both prokaryotic and eukaryotic organisms.

关键词： eYGFPuv green fluorescent protein screening marker

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Identification and Function of a Novel Gene,ORF188,Associated with Cytoplasmic Effects on Oil Content in Brassica napus

Identification and Function of a Novel Gene,ORF188,Associate...

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2014年中国作物学会学术年会

作者： Wanjun Hao Wei Hua shihang fan Xinfa Wang Zhiyong Hu Jing Liu Hongli Yang Hanzhong Wang Key Laboratory for Biological Sciences of Oil Crops Ministry of AgricultureOil Crops Research InstituteChinese Academy of Agriculture Sciences College of Life Sciences Hubei University

Seed oil content(SOG) is an important agronomic trait in rapeseed(Brassica napus L.).Previous research showed cytoplasmic effect playing an important role on SOC in ***,the underlying molecular mechanisms remain largely *** this study,we identified and characterized a novel gene,ORF188,associated with cytoplasmic effect on SOC in *** on previous study,we obtained two rapeseed inbred lines 51218 and 56366,between which cytoplasmic effect had significant difference and reached seven per cent on *** understand the genetic bases,we sequenced plastid and mitochondrial genomes of the two rapeseed *** compared their plastid genomes and found many small genetic *** plastid genome,there were many small genetic variations between their mitochondrial *** surprisingly,it seemed rearrangements had happened between the two mitochondrial *** a result,there are fourteen different ORFs which are specific to one of the two mitochondrial ***,we conjectured that these fourteen specific ORFs are the reasons for different cytoplasmic effect on SOC between 51218 and *** further validate the conjecture,we compared the transcriptomics of reciprocal cross F which were made between 51218 and *** expected,there were only five cytoplasmic genes with differential ***,all these five genes were specific mitochondrial ***,this result validated our conjecture and reduced the candidate gene number to *** another way,a set of 48 rapeseed inbred lines possessing different types of cytoplasm were used for association *** measure cytoplasmic effect of the 48 rapeseed lines,reciprocal cross were made between 56366 and each one of *** cytoplasmic effects were identified as the difference of SOC between reciprocal cross ***,fourteen specific mitochondrial ORFs were designed as molecular marker and used for *** significant associations for cytoplasmic effect were d

关键词： Brassica napus cytoplasmic effect seed oil content ORF188 ATP

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Effective method for isolating liquid endosperm and improved procedure for subsequent DNA extraction in Brassica napus

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Oil Crop Science 2016年第2期1卷 20-25页

作者： Jun LI shihang fan Xingchao SUN Zhiyong HU Jing LIU Xiaoyi ZHU Ming ZHENG Fengming SUN Hongli YANG Liang ZHANG Linbin DENG Hongfang LIU Wei HUA Key Laboratory of Biology and Genetic Improvemtent of Oil Crops Ministry of AgricultureOil Crops Research InstituteChinese Academy of Agriculture Sciences

The important agronomic traits of seed size and seed oil content in oilseed rape were af-fected by maternal effect. The maternal tissue endosperm, generated by fertilization of the diploid central cell in the female gametophyte, is essential for embryo and seed development. Imprinting is primarily re-stricted to the endosperm of flowering plants and is associated with differential DNA methylation of paren-tal alleles. Therefore, it is necessary to find out whether the endosperm influences these traits. If so, whether DNA methylation participated in these processes？ To answer these questions, isolation of pure liquid endosperm qualified for extraction genomic DNA and RNA was the prerequisite. However, it was not easy to conduct this isolation due to endosperm is encapsulated by tiny seed coat and it adjoins to de-veloping embryo. This research adopted a procedure for isolation pure lipid endosperm from 25DAF （days after flowering） ovules of Brassica napus and improved procedure for subsequent DNA extraction. With the optimized procedures, we successfully extracted genomic DNA of liquid endosperm from 500 mg o-vules with high concentration （ ≥2. 5μg/μl, 200μl） and high purity, which were proved by the subse-quent analysis.

关键词： rapeseed liquid endosperm genomic DNA effective

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Overexpression of Sddt enhances drought tolerance in Arabidopsis and Brassica napus by regulating micro RNA activity

Overexpression of Sddt enhances drought tolerance in Arabido...

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2016年全国植物生物学大会

作者： Zhiyong Hu Liang Zhang Jing Liu Gaomiao Zhan Hongli Yang Xingchao Sun Linbin Deng shihang fan Hanzhong Wang Wei Hua Oil Crops Research Institute of the Chinese Academy of Agricultural Sciences Key Laboratory of Biology and Genetic Improvement of Oil CropsMinistry of Agriculture

In plants,high root density and larger root surface area can be maintained or expanded to promote water uptake,a process that plays crucial role in drought stress tolerance. In our study,a gene named Sddt(a semi-dwarfism and drought tolerance gene) was found to regulate the development of lateral root growth in Arabidopsis and Brassica napus. Compared with wild type,transgenic plants overexpressing Sddt showed increased lateral roots. Using yeast-2-hybrid,we demonstrated that Sddt interacted with DCL1. Hybridization of Sddt-overexpressing plants with miRNA activity reporter plants indicated that Sddt greatly enhanced miRNA activity. Furthermore,by small RNA sequencing,we identified 96 known differentially expressed miRNA genes. Among them,the miRNA160 family was up-regulated and its target genes,ARF10,-16,and-17,were down-regulated. Our results suggest that the overexpression of Sddt enhances micro RNA activity via its interaction with DCL1,decreases expression of ARF genes through miRNA160,promoting both lateral root development and increased drought stress tolerance. These findings will help us to better understand the mechanism of root development during drought stress,and implies that Sddt has potential applications in genetic engineering to improve crop tolerance.

关键词： Arabidopsis Brassica napus drought tolerance miRNA activity Sddt miRNA160

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甘蓝型油菜线粒体功能未知基因ORF117的载体构建与转化

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氨基酸和生物资源 2015年第2期37卷 43-51页

作者：范世航郝万军华玮陈永勤杨之帆湖北大学生命科学学院湖北武汉430062 中国农业科学院油料作物研究所/农业部油料作物生物学与遗传育种重点实验室湖北武汉430062

本研究克隆了甘蓝型油菜线粒体功能未知基因ORF117,构建了带有线粒体转运信号肽序列(TP)的植物过表达载体35S∷TP-ORF117、35S∷TP-EGFP和35S∷TP-ORF117-EGFP,转化野生型拟南芥并进行抗除草剂筛选,共获得纯合的转基因拟南芥株系62个。q... 详细信息

本研究克隆了甘蓝型油菜线粒体功能未知基因ORF117,构建了带有线粒体转运信号肽序列(TP)的植物过表达载体35S∷TP-ORF117、35S∷TP-EGFP和35S∷TP-ORF117-EGFP,转化野生型拟南芥并进行抗除草剂筛选,共获得纯合的转基因拟南芥株系62个。qPCR表明,ORF117在转基因材料中得到高效过表达。用转35S∷TP-EGFP、35S∷TPORF117-EGFP拟南芥制备原生质体,经线粒体特异染料染色,在激光共聚焦显微镜下做线粒体、GFP共定位检测,GFP蛋白和ORF117-EGFP融合蛋白被准确定位到了线粒体。

关键词：甘蓝型油菜线粒体转运肽载体构建

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Tetra-color superresolution microscopy based on excitation spectral demixing

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Light(Science & Applications) 2023年第1期12卷 99-108页

作者： Wanyan Wu shihang Luo Chunyan fan Tianjie Yang Shuwen Zhang Wenxiang Meng Tao Xu Wei Ji Lusheng Gu Institute of Biophysics Chinese Academy of SciencesBeijing 100101China College of Life Science University of Chinese Academy of SciencesBeijing 100049China State Key Laboratory of Molecular Developmental Biology Institute of Genetics and Developmental BiologyInnovation Academy for Seed DesignChinese Academy of SciencesBeijing 100101China School of Future Technology University of Chinese Academy of SciencesBeijing 100049China College of Advanced Agricultural Sciences University of Chinese Academy of SciencesBeijing 100049China Bioland Laboratory Guangzhou 510005China Guangzhou Laboratory Guangzhou 510030China

Multicolor imaging allows protein colocalizations and organelle interactions to be studied in biological research,which is especially important for single-molecule localization microscopy(SMLM).Here,we propose a multicolor method called excitation-resolved stochastic optical reconstruction microscopy(ExR-STORM).The method,which is based on the excitation spectrum of fluorescent dyes,successfully separated four spectrally very close far-red organic fluorophores utilizing three excitation lasers with cross-talk of less than 3%.Dyes that are only 5 nm apart in the emission spectrum were resolved,resulting in negligible chromatic *** method was extended to three-dimensional(3D)imaging by combining the astigmatic method,providing a powerful tool for resolving 3D morphologies at the nanoscale.

关键词： excitation spectral mixing

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孔隙率对环形阵列圆柱结构局部冲刷的影响

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上海交通大学学报 2022年第5期56卷 664-674页

作者：刘诗航娄晓帆唐国强王滨大连理工大学海岸和近海工程国家重点实验室辽宁大连116024 浙江省深远海风电技术研究重点实验室杭州311122

通过物理实验研究了群桩结构在单向流作用下的局部冲刷问题,重点关注群桩结构孔隙率对局部冲刷的影响作用.物理实验设置中,在群桩模型外直径固定的条件下,通过改变单桩的数量改变群桩结构的孔隙率,主要研究群桩孔隙率对局部冲刷平衡深... 详细信息

通过物理实验研究了群桩结构在单向流作用下的局部冲刷问题,重点关注群桩结构孔隙率对局部冲刷的影响作用.物理实验设置中,在群桩模型外直径固定的条件下,通过改变单桩的数量改变群桩结构的孔隙率,主要研究群桩孔隙率对局部冲刷平衡深度、范围及时间尺度的影响作用.结果表明:模型的孔隙率对群桩周围的冲刷有着显著的影响作用.当孔隙率p>0.7时,冲刷主要由单元桩周围的冲刷决定;当p<0.5时,冲刷趋向于与群桩外直径相等的单桩冲刷的模式;当0.5

关键词：局部冲刷单向流群桩孔隙率

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振动条件下铣刀切削过程物质信息流构建方法

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振动与冲击 2024年第20期43卷 200-212,262页

作者：范丽丽姜彬赵培轶李世航哈尔滨理工大学机械动力工程学院哈尔滨150080

振动条件下铣刀切削过程物质的传递转换关系是揭示其加工表面形成过程多时变性的关键。依据立铣刀、刀齿、工件之间物质能量的输入输出关系与立铣刀动态切削行为特性,构建铣刀切削过程的物质流结构,揭示各组元间的传递转换关系;利用铣... 详细信息

振动条件下铣刀切削过程物质的传递转换关系是揭示其加工表面形成过程多时变性的关键。依据立铣刀、刀齿、工件之间物质能量的输入输出关系与立铣刀动态切削行为特性,构建铣刀切削过程的物质流结构,揭示各组元间的传递转换关系;利用铣刀、工件等不同物质载体的结构、运动及切削力等参数,定量表征物质流结构各节点的瞬时切削状态;构建物质流结构相邻节点的邻接矩阵,阐明相邻节点之间的传递关系;采用均方根值、峭度、主频与分形维数,揭示铣刀切削过程中物质信息流内部节点的动态特性。对物质信息流输入端、切削层节点及输出端进行试验验证。结果表明,该方法可揭示机床驱动下铣刀切削过程物质的传递与转换过程,为铣削工艺设计提供基础模型。

关键词：铣削振动铣刀物质流物质信息流流结构

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微波改性膨润土对含磷废水的吸附性能研究

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非金属矿 2014年第1期37卷 69-71页

作者：范恒聂锦旭吴光锋刘世杭广东工业大学土木与交通工程学院广东广州510006

以C8烷基多糖苷季铵盐和AlCl3为改性剂对钠基膨润土进行改性,制备改性膨润土,利用其处理含磷废水,考察了微波辐射功率、时间、改性剂用量、膨润土投加量、pH值、磷初始质量浓度对磷去除效果的影响。结果表明,在微波辐射功率为320 W,时间... 详细信息

以C8烷基多糖苷季铵盐和AlCl3为改性剂对钠基膨润土进行改性,制备改性膨润土,利用其处理含磷废水,考察了微波辐射功率、时间、改性剂用量、膨润土投加量、pH值、磷初始质量浓度对磷去除效果的影响。结果表明,在微波辐射功率为320 W,时间为10 min,有机改性剂用量为3.0 g/100g,铝改性剂用量为9 mmol/g条件下制备的改性膨润土,其投加量为12 g/L,废水pH值为6时,对初始质量浓度50 mg/L的模拟含磷废水去除率达到96%以上,对实际含磷矿山废水平均去除率为87.83%。

关键词：微波改性膨润土吸附磷

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