苦荞突变体库构建与突变体中芦丁合成相关基因表达分析

苦荞是重要的小杂粮作物之一,营养物质丰富,是天然芦丁的重要来源。突破苦荞育种难题,创制苦荞新种质是目前研究的重要方面。本试验利用甲基磺酸乙酯(EMS)构建了黑丰1号苦荞突变体库,明确了当EMS浓度为1.2%时,诱变效果较好。通过对M1突变株表型观察统计,共获得叶色、叶型、株型、粒型变异单株102株,突变率为3.85%;高效液相色谱技术(HPLC)测定1000株M3材料,获得高芦丁含量突变株系2个和低芦丁突变体株系5个;qRT-PCR对芦丁含量突变体株系中芦丁代谢关键酶基因(CHS、F3H、4CL、FLS、UFGT)进行表达量分析,发现不同株系中上述基因的表达量与芦丁含量相关性不明显,但个别基因如FtFLS基因表达量在高芦丁含量突变体中达到对照的4.55倍。通过突变体的筛选丰富了苦荞基因资源,创新了苦荞新种质,也为苦荞芦丁代谢的分子基础研究提供了材料保证与技术支持。     

生物反应器与药用植物毛状根的大规模培养

利用药用植物毛状根培养生产次生代谢产物具有极大的生产潜力,而开发适合毛状根培养的反应器,又是毛状根生产天然产物工业化的关键。本文系统地介绍了各种用于毛状根培养的生物反应器,对通气搅拌式、气升式、超声雾化式等生物反应器各自的特点及优势进行了详细阐述,进一步讨论了各种反应器对于毛状根生长和次生代谢物积累的影响,并提出药用植物毛状根大规模培养的生物反应器在今后的发展方向。     

苦荞地方种质资源的遗传多样性分析

利用SSR分子标记技术对中国苦荞主产区陕西、云南、四川、西藏等地的82份苦荞地方种质的遗传多样性进行了分析,以揭示中国特有的作物种质--苦荞地方种质资源的遗传多样性,促进苦荞优良品种的选育.结果显示:(1)所用25个SSR引物中有13个引物在苦荞地方品种中具有多态性,且扩增条带的稳定性较好,共扩增出208条条带,其中多态性条带200条,占总数的96.2%;(2)聚类分析结果显示,82份苦荞材料的遗传相似系数(GS)分布于0.52～0.85之间,平均值为0.69,在GS值为0.722的水平上,82份材料被聚为10大类群.研究表明82份苦荞品种间遗传多样性明显,具有丰富的遗传基础.     

野葛毛状根离体培养与异黄酮生产

用发根农杆菌（Agrobacterium rhizogenes）R1601菌株感染野葛（Pueraria lobata(Willd.)Ohwi）叶片外植体后获得的毛状根,经连续4次继代培养,鲜重增加到最初的6.2倍。PH为5.5的1/2SH液体培养基较适合野葛毛状根的生长。与自然根相比,毛状根在无激素的1/2SH液体培养基中离体培养20d后,鲜重增加22.6倍,释放入培养液中的异黄酮量增加10.6倍。毛状根生产异黄酮的量略高于自然根,且根中含量明显高于愈伤组织及茎中异黄酮的含量。培养液体积影响毛状根的离体生长及异黄酮的释放。     

毛蕊花和粟米草毛状根培养系统的建立(简报)

用发根农杆菌Ａ４、Ｒ１６０１、ＡＴＣＣ１５８３４从毛蕊花和粟米草的无菌子叶中诱导出毛状根,经高压纸电泳鉴定,这两种毛状根中均含有甘露碱,并测定了它们的生长曲线。     

顶空气相色谱法检测苦荞麦总黄酮提取物的有机溶剂残留量

为控制使用AB-8型大孔树脂后苦荞麦总黄酮提取物的质量,采用顶空气相色谱法,对提取物中正己烷、苯、甲苯、对二甲苯、间二甲苯、邻二甲苯、乙苯和苯乙烯等8种残留物进行检测。精密度RSD均小于6％,峰面积与浓度均有良好的线性关系,最低检测限为0．016—0．0640g／mL,回收率均符合要求。试验结果表明,方法稳定、操作简便、准确可靠,适用于使用AB-8型大孔树脂后苦荞麦总黄酮提取物中有机溶剂残留量的检测。     

苦荞NAC转录因子FtNAC17的鉴定及表达分析

植物特有的NAC转录因子参与植物生长发育和逆境响应。研究苦荞中NAC转录因子在非生物胁迫中的应答,为阐明基因功能提供理论依据。以苦荞为试验材料,克隆一个NAC家族基因,对其进行生物信息学分析。通过荧光定量PCR技术检测该基因在干旱、低温、盐、茉莉酸甲酯、脱落酸和赤霉素胁迫时的响应情况。该基因编码272个氨基酸,将其命名为FtNAC17,GenBank登录号为MT641452。基因结构分析表明,FtNAC17由2个外显子和1个内含子组成。氨基酸序列多重比对和进化关系分析表明,FtNAC17蛋白与拟南芥ANAC002亲缘关系最近,属于NAC转录因子家族中同一亚组。FtNAC17对干旱、低温、盐等非生物胁迫均有不同程度的响应。     

Indole Glucosinolate Biosynthesis Limits Phenylpropanoid Accumulation in Arabidopsis thaliana

Plants produce an array of metabolites (including lignin monomers and soluble UV-protective metabolites) from phenylalanine through the phenylpropanoid biosynthetic pathway. A subset of plants, including many related to Arabidopsis thaliana, synthesizes glucosinolates, nitrogen- and sulfur-containing secondary metabolites that serve as components of a plant defense system that deters herbivores and pathogens. Here, we report that the Arabidopsis thaliana reduced epidermal fluorescence5 (ref5-1) mutant, identified in a screen for plants with defects in soluble phenylpropanoid accumulation, has a missense mutation in CYP83B1 and displays defects in glucosinolate biosynthesis and in phenylpropanoid accumulation. CYP79B2 and CYP79B3 are responsible for the production of the CYP83B1 substrate indole-3-acetaldoxime (IAOx), and we found that the phenylpropanoid content of cyp79b2 cyp79b3 and ref5-1 cyp79b2 cyp79b3 plants is increased compared with the wild type. These data suggest that levels of IAOx or a subsequent metabolite negatively influence phenylpropanoid accumulation in ref5 and more importantly that this crosstalk is relevant in the wild type. Additional biochemical and genetic evidence indicates that this inhibition impacts the early steps of the phenylpropanoid biosynthetic pathway and restoration of phenylpropanoid accumulation in a ref5-1 med5a/b triple mutant suggests that the function of the Mediator complex is required for the crosstalk.     

Complete Chloroplast Genome Sequence of Tartary Buckwheat (Fagopyrum tataricum) and Comparative Analysis with Common Buckwheat (F. esculentum)

We report the chloroplast (cp) genome sequence of tartary buckwheat (Fagopyrum tataricum) obtained by next-generation sequencing technology and compared this with the previously reported common buckwheat (F. esculentum ssp. ancestrale) cp genome. The cp genome of F. tataricum has a total sequence length of 159,272 bp, which is 327 bp shorter than the common buckwheat cp genome. The cp gene content, order, and orientation are similar to those of common buckwheat, but with some structural variation at tandem and palindromic repeat frequencies and junction areas. A total of seven InDels (around 100 bp) were found within the intergenic sequences and the ycf1 gene. Copy number variation of the 21-bp tandem repeat varied in F. tataricum (four repeats) and F. esculentum (one repeat), and the InDel of the ycf1 gene was 63 bp long. Nucleotide and amino acid have highly conserved coding sequence with about 98% homology and four genes—rpoC2, ycf3, accD, and clpP—have high synonymous (Ks) value. PCR based InDel markers were applied to diverse genetic resources of F. tataricum and F. esculentum, and the amplicon size was identical to that expected in silico. Therefore, these InDel markers are informative biomarkers to practically distinguish raw or processed buckwheat products derived from F. tataricum and F. esculentum.     

苦荞提取物磷脂复合物的制备工艺研究

本文以苦荞总黄酮与大豆磷脂的复合率为评价指标,采用单因素考察对苦荞提取物与大豆磷脂形成复合物的制备工艺技术条件进行系统研究,并测定复合物在正辛醇和不同pH水溶液中的表观油水分配系数。结果表明:筛选出的最佳工艺条件为,以无水乙醇为反应溶剂,提取物的浓度为1 mg/mL,提取物与磷脂的质量比为1∶2,反应温度为40℃,反应时间24 h。与原提取物比较,磷脂复合物中总黄酮的表观油水分配系数有了显著的提高。按照该制备工艺制备的复合物复合率达到80%以上,同时结果表明磷脂复合物明显改善了苦荞总黄酮的脂溶性。     

利用气升式内环流生物反应器培养青蒿毛状根生产青蒿素

利用自制的气升式内环流生物反应器进行青蒿(Artemisia annua L.)毛状根多层培养生产青蒿素。毛状根培养物在培养过程中均匀分布在生物反应器的筛网间,或以不锈钢网为附着点向四周生长,在25 ℃和12 h/d光照周期下,经20 d分批培养获得生物量干重22.57 g/L,青蒿素产量374.4 mg/L,并对培养过程中蔗糖、磷酸盐、硝酸盐和氨盐消耗的动力学进行了分析。     

利用气升式内环流生物反应器培养青蒿毛状根生产青蒿素

利用自制的气升式内环流生物反应器进行青蒿（ArtemisiaannuaL．）毛状根多层培养生产青蒿素。毛状根培养物在培养过程中均匀分布在生物反应器的筛网间，或以不锈钢网为附着点向四周生长，在25℃和12h／d光照周期下，经20d分批培养获得生物量干重22．57g／L，青蒿素产量374．4mg／L，并对培养过程中蔗糖、磷酸盐、硝酸盐和氨盐消耗的动力学进行了分析。     

微波提取苦荞麦麸皮总黄酮工艺研究

本文对苦荞麦麸皮总黄酮微波提取工艺进行了研究。试验结果表明:微波提取的最佳工艺条件为微波功率中档,微波加热120 s,乙醇浓度80%,料液比1:50,该工艺条件下总黄酮得率达5.51%;与传统提取方法相比,微波提取法具有节省时间、节约能量、提取效率高、控制方便等优点。     

2-Geranyl-1,4-naphthoquinone,a Possible Intermediate of Anthraquinones in a Sesamum indicum Hairy Root Culture

2-Geranyl-1,4-naphthoquinone was isolated from the hairy root culture of Sesamum indicum. The structure was determined to be 2-[(E)-3,7-dimethylocta-2,6-dienyl]-1,4-naphthoquinone on the basis of spectroscopic evidence and chemical synthesis. The production of anthrasesamones A, B and C by the hairy root culture was also confirmed for the first time.     

Tissue-Autonomous Phenylpropanoid Production Is Essential for Establishment of Root Barriers

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西洋参发根的诱导及不同外源物质对发根生长和皂苷含量的影响

利用发根农杆菌A4菌株在西洋参根外植体上直接诱导产生发根,并分析不同浓度的植物激素6-BA、NAA,前体物醋酸镁(Mg(Ac)2)、L-亮氨酸和诱导子茉莉酸甲酯(MJA)、水杨酸(SA)对西洋参(Panax quinquefolium L.)发根生长和皂苷含量的影响。同时,研究MJA与SA组合作用对皂苷含量的影响。采用高效液相色谱法测定发根中单体皂苷Rb1的含量。结果显示,适宜浓度的外源物质均可促进皂苷含量的增加,MJA尤其明显。     

苦荞类过敏原的原核表达及多克隆抗体制备

以苦荞种子发育期全长cDNA文库中获得的与甜荞16 kD过敏原同源的基因BALP(Buckwheat Allergenlike protein,BALP; GenBank登录号GO496294)为基础,构建了原核表达载体pET47b-BALP,实现了在大肠杆菌Rosetta gami2 (DE3)中的高效表达,并对诱导...