Antioxidant Activity of Ethyl Acetate Extract of Amomum villosum Lour.

[Objectives]To study the antioxidant activity of the ethyl acetate extract of Amomum villosum Lour.[Methods]The removal rate of chelated iron ions,hydroxyl radicals,superoxide anion free radicals and DPPH free radicals by the ethyl acetate extract of A.villosum Lour.was determined by UV spectrophotometer.[Results]0.5000μg/mL ethyl acetate extract of A.villosum Lour.had the strongest ability to chelate with ferrous ions and to remove hydroxyl radicals,superoxide anion free radicals and DPPH free radicals.The ability to chelate with ferrous ions was 95.14%,and the removal rate of the above free radicals was 86.217%,81.44%,and 85.16%.[Conclusions]The ethyl acetate extract of A.villosum Lour.had a strong antioxidant effect,and its antioxidant capacity was related to the sample concentration,which provides a theoretical basis for its application in the development of antioxidant skin care products.

簇毛麦(Dasypyrum villosum)GA20ox基因的分离和鉴定

通过RT-PCR技术克隆了簇毛麦(Dasypyrum villosum)赤霉素生物合成关键酶GA20ox基因的全长序列,命名为DvGA20ox,GenBank登录号为EU142949.该基因全长1080bp,推测编码359个氨基酸的蛋白质,具有植物GA20ox基因的典型保守结构域LPWKET和NYYPXCQKP.该基因推导编码蛋白质与小麦、大麦和黑麦草GA20ox蛋白的同源性分别为98%、97%和91%.该序列重组到原核表达载体pET-32a(+)获得的重组子pET-32a(+)-DvGA20ox转化大肠杆菌BL21pLysS后,IPTG诱导表达,SDS-PAGE分析表明,DvGA20ox基因在大肠杆菌中获得了高效表达,融合蛋白相对分子质量为55×103,与理论值相符.结果为深入研究DvGA20ox蛋白的结构与功能以及该基因与株高发育的关系奠定了基础.

阳春砂(Amomum villosum Lour.)农家种热科2性状初报

【目的】阳春砂(Amomum villosum Lour.)是砂仁中的佳品,具有悠久的药用历史。通过系统分析阳春砂农家种热科2特征性状,为阳春砂品种识别和生产种植提供依据。【方法】对阳春砂农家种热科2植株、花、果实等性状进行跟踪调查研究与对比分析。【结果】热科2阳春砂植株高大粗壮,株高2.84 m,茎粗13.12 mm;叶片大而厚,叶长43.52 cm、宽6.46 cm,外侧叶缘波状不明显、叶面具平行于主脉的突起线,叶舌长椭圆形、棕色、长0.95 mm;花序“曲线型”,花梗长9.70 mm,小花数9.3朵,唇瓣长2.24 mm、宽2.39 mm,中脉黄色而染紫红色、顶端黄色外延、基部紫红色“八”字型痂状斑;果实扁圆形、褐色,内含种子23.2粒,千粒质量14.14 g,种子团挥发油体积分数3.6%;花期和果实成熟期晚。【结论】明确了热科2阳春砂的形态特征性状,可作为砂仁DUS测试指南的标准品种,为该品种的精准识别以及阳春砂优异资源保护与选育种奠定基础。

簇毛麦(Dasypyrumvillosum)矮秆突变体的鉴定

为给簇毛麦杂交后代群体中自然突变产生的矮秆突变体在小麦遗传改良中合理有效的利用提供理论依据,对该矮秆突变体的生物学及遗传学特性进行了初步研究。该突变体在苗期与对照植株无明显差异,在成熟期突变体株高30 cm,约为对照株高的1/3。突变株茎的倒一、二节间(最上部的两个茎节)长度分别为对照植株长度的1/9和1/6。细胞学观察表明,突变体的茎节细胞纵向延长受到抑制。此外,该突变株还表现出部分雄性不育,自然结实率低。花粉活力检测发现,突变株可育花粉只有20%,在外施GA3后花粉育性可得到恢复,推测该突变体为赤霉素敏感型矮秆突变体。遗传分析表明,矮秆突变性状受一个隐性基因控制。

阳春砂(Amomum villosum Lour.)农家栽培种‘热科1’特征性状初报

阳春砂(Amomum villosum Lour.)是我国四大南药之一,具有极高的药用价值。我国阳春砂栽培种源混杂、退化严重等问题制约着其品质与药效的提升,亟待加强优质资源的筛选与改良。本研究通过调查阳春砂农家栽培种‘热科1’植物学性状,发现:阳春砂主茎高度208㎝,叶缘波状程度强且整齐,叶舌椭圆形、棕色,叶鞘绿黄色、边缘轻度棕色带;新抽芽被鲜艳棕红色膜质鳞片;花序"L"型、花梗长8.46㎝,小花数11.9朵;唇瓣中脉黄染深紫红,花粉囊覆盖处中部黄色、边缘染微红色;果实椭圆形、基部微三棱形,种子团挥发油含量3.7%,乙酸龙脑酯含量1.68%;种子褐色、不规则形状,千粒重11.42 g。本研究对阳春砂农家栽培种‘热科1’植株、花、果实与种子特性进行了详细的描述,具有鲜明的特征,作为砂仁DUS测试指南的标准品种,为识别‘热科1’品种提供科学依据,为砂仁品种保护奠定基础。

簇毛麦(Dasypyrum villosum L.)中一个γ-醇溶蛋白基因序列的研究

目的:研究簇毛麦中一个新的γ-醇溶蛋白基因序列及结构特点,为小麦品质育种和该类基因的进化分析提供资料。方法:利用基因组PCR技术从簇毛麦基因组中克隆到1个γ-醇溶蛋白基因(Dv-γ)的全编码序列,利用生物信息学研究其序列结构特点。结果:该序列与已知的γ-醇溶蛋白有着很高的相似性。推导的氨基酸序列包含5个典型的结构域,其中含有8个保守半胱氨酸残基,高变重复区Ⅱ中的重复单元与其他物种来源基因有较大区别。γ-醇溶蛋白中常见的乳糜泄抗原决定簇在其内部也有发现。进化分析表明,此基因与亚家族Ⅰ中的γ-醇溶蛋白基因有着较近的亲缘关系。结论:获得了一个新的γ-醇溶蛋白基因。

Genetic behavior of Triticum aestivum–Dasypyrum villosum translocation chromosomes T6V#4S·6DL and T6V#2S·6AL carrying powdery mildew resistance

T6V#2S·6AL and T6V#4S·6DL translocation chromosomes developed from the cross of wheat and different Dasypyrum villosum accessions have good powdery mildew （PM） resistance, but their pairing and pyramiding behavior remains unclear. Results in this study indicated that the pairing frequency rate of the two differently originated 6VS chromosomes in their F1 hybrid was 18.9% according to genomic in situ hybridization （GISH）; the PM resistance plants in the F2 generation from the cross between T6V#4S·6DL translocation line Pm97033 and its PM susceptible wheat variety Wan7107 was fewer than expected. However, the ratio of the resistant vs. the susceptible plants of 15：1 in the F2 generation derived from the cross between the two translocation lines of T6V#2S·6AL and T6V#4S·6DL fitted well. Plants segregation ratio （homozygous：heterozygous：lacking） revealed by molecular marker for T6V#4S·6DL or T6V#2S·6AL in their F2 populations fitted the expected values of 1：2：1 well, inferring that the pairing of the two alien chromosome arms facilitates the transmission of T6V#4S·6DL from the F1 to the F2 generation. A quadrivalent was also observed in 21% of pollen mother cells （PMCs） of homozygote plants containing the two pairs of translocated chromosomes. The chromosome pairing between 6V#2S and 6V#4S indicates that it will be possible to obtain recombinants and clarify if the PM resistance determinant on one alien chromosome arm is different from that on the other.

Pm67, a new powdery mildew resistance gene transferred from Dasypyrum villosum chromosome 1V to common wheat(Triticum aestivum L.)

Powdery mildew, caused by the biotrophic fungus Blumeria graminis f. sp. tritici(Bgt), is a global disease that poses a serious threat to wheat production. To explore additional resistance gene, a wheatDasypyrum villosum 1 V#5(1 D) disomic substitution line NAU1813(2 n = 42) with high level of seedling resistance to powdery mildew was used to generate the recombination between chromosomes 1 V#5 and1 D. Four introgression lines, including t1 VS#5 ditelosomic addition line NAU1815, t1 VL#5 ditelosomic addition line NAU1816, homozygous T1 DL·1 VS#5 translocation line NAU1817, and homozygous T1 DS·1 VL#5 translocation line NAU1818 were developed from the selfing progenies of 1 V#5 and 1 D double monosomic line that derived from F1 hybrids of NAU1813/NAU0686. All of them were characterized by fluorescence in situ hybridization, genomic in situ hybridization, 1 V-specific markers analysis, and powdery mildew tests at different developmental stages. A new powdery mildew resistance gene named Pm67 was physically located in the terminal bin(FL 0.70–1.00) of 1 VS#5. Lines with Pm67 exhibited seedling stage immunity and tissue-differentiated reactions at adult plant stage. The sheaths, stems, and spikes of the Pm67 line were still immune, but the leaves showed a low degree of susceptibility.Microscopic observation showed that most penetration attempts were stopped in association with papillae on the sheath, and colonies cannot form conidia on the susceptible leaf of Pm67 line at adult plant stage, suggesting that the defence layers of the Pm67 line is tissue-differentiated. Thus, the T1 DL·1 VS#5 translocation line NAU1817 provides a new germplasm in wheat breeding for improvement of powdery mildew resistance.

Development of EST-PCR markers specific to the long arm of chromosome 6V of Dasypyrum villosum

Expressed sequence tags-derived polymerase chain reaction(EST-PCR) molecular markers specific for alien chromosomes can be used to not only monitor the introgressed alien chromatin in wheat background, but also provide the evidence of the syntenic relationship between homoeologous chromosomes. In the present study, in order to develop high density and evenly distributed molecular markers specific for chromosome 6 VL of Dasypyrum villosum, 297 primer pairs were designed based on the expressed sequence tags(EST) sequences, which were previously mapped in different bins of the long arms of wheat homoeologous 6 AL, 6 BL, and 6 DL. By using the Triticum aestivum, D. villosum, T. durum-D. villosum amphiploid, and T. aestivum-D. villosum alien chromosome lines involving chromosome 6 V, it was found that 32(10.77%) primers could amplify specific bands for chromosome 6 V, and 31 could be allocated to chromosome arm 6 VL. These 6 VL specific markers provided efficient tools for the characterization of structural variation involving the chromosome 6 VL in common wheat background as well as for the selection of useful genes located on 6 VL in breeding programs.

春砂仁(Amomum villosum Lour.)种子发芽的试验研究

春砂仁种植于广东、广西、福建、云南、四川等省,以广东为主要产区.生产中多采用分株繁殖的方法,由于种苗不足,大面积发展受到限制.近年不少产区试用种子繁殖,又往往因采收、贮藏、播种或长途运输等处理不当,致使种子发芽率很低.为了探讨春砂仁种子发芽的特性及其与环境之间的关系,从而找出催芽的有效措施,提高种子发芽率,我们从197s年开始,采用几种方法进行了种子发芽的比较试验.

Introgression of sharp eyespot resistance from Dasypyrum villosum chromosome 2VL into bread wheat

Wheat sharp eyespot, a stem disease caused by the soilborne fungus Rhizoctonia cerealis van der Hoeven,has become a threat to wheat production worldwide. Exploiting resistance resources from wild relatives of wheat is a promising strategy for controlling this disease. In this study, a new wheat–Dasypyrum villosum T2DS·2V#4L translocation line in the background of Chinese Spring(CS) showed stable resistance to R. cerealis. Introgression of the T2DS·2V#4L chromosome into wheat cultivar Aikang 58 by backcrossing produced a marked increase in sharp eyespot resistance in NIL-T2DS·2V#4L in comparison with NILT2DS·2DL, and no detrimental effects of 2V#4L on agronomic traits were observed in the BC2F2, BC2F2:3,and BC2F2:4generations. Flow-sorted sequencing of 2V#4L yielded 384.3 Mb of assembled sequence, and8836 genes were predicted of which 6154 had orthologs in at least one of the 2AL, 2BL, and 2DL arms of CS, whereas 1549 genes were unique to 2V#4L. About 100,000 SNPs were detected in genes of 2V#4L and2DL in 10 sequenced bread wheat cultivars. A Kompetitive Allele Specific Polymerase chain reaction and30 conserved ortholog sequence markers were developed to trace the 2V#4L chromatin in wheat backgrounds. T2DS·2V#4L compensating translocation lines represent novel germplasm with sharp eyespot resistance and the markers will allow rapid detection in breeding programs.

Characterization and Expression Profiling of Genes Encoding the Gibberellin 3-oxidase in Dasypyrum villosum Dwarf Mutant

Gibberellin 3-oxidase catalyzes the conversion of inactive gibberellin(GA) species into GAs with biological activity and it is subjected to strict developmental controls in the life cycle of a plant. In this study, 33 gene sequences, encoding the gibberellin 3-oxidase(GA3ox) from Dasypyrum villosum and its dwarf mutant, were obtained. Each contained a 1 107 bp coding sequence(CDS) that encoded a putative protein containing 369 amino acids. The GA3ox protein showed 77% to 97% homology and shared the major conserved structural domains of GA3ox proteins with rice, sorghum bicolor, oat, barley, and wheat. Sequence alignment showed that there were 20 single nucleotide polymorphisms(SNPs) and 22 Insertion/deletions(In Dels) among these sequences, which could be divided into 2 haplotypes, haplotypes Ⅰ and Ⅱ. Haplotype Ⅰ was found in the wild type and was1 495 bp in length, and haplotype Ⅱ was found in the dwarf mutant and was 1 485 bp in length. The Q-PCR results showed that GA3ox was expressed in the leaves, roots, internodes, and stem nodes, and that there was a significant difference in the transcript level of the GA3ox between the wild type and dwarf mutant. The transcript levels of GA3ox in the leaves at the seedling stage, stem elongation stage and the heading stage, in the root and stem nodes at the stem elongation stage and in the internodes at the heading stage of the wild type, were significantly higher than those in the dwarf mutant. However, GA3ox expression in the rest of the wild type tissues at the 3 stages was slightly higher than or not different from the dwarf mutant.The results suggested that the wild type and mutant allele sequences of GA3ox in D. villosum showed 2 amino acid changes in exons and variations in the lengths of introns or the SNPs in introns, which most probably impaired the function of the enzyme,affected the GA3ox expression level, and eventually gave rise to dwarfing.

毛姜花中半日花烷型二萜的分离与鉴定

毛姜花(Hedychium villosum Wall)根茎粉碎后用工业甲醇浸提,浸提物用乙酸乙酯-水萃取。采用正反相硅胶柱层析、葡聚糖凝胶柱层析和HPLC层析等分离技术,通过NMR等方法对乙酸乙酯浸膏中的化学成分进行分离鉴定,得到5个半日花烷型二萜化合物,分别是hedychenone(1)、yunnancoronarin D(2)、coronarin A(3)、yunnancoronarin A(4)和yunnancoronarin E(5),均为首次从毛姜花中分离得到。

一个成株抗白粉病小麦-簇毛麦T2DS·2V#6L易位系的选育

白粉病是威胁全球小麦生产的一种严重病害,从小麦野生近缘种发掘成株抗白粉病基因资源对培育持久抗性品种十分重要。本研究从硬粒小麦-簇毛麦01I139(V#6)双二倍体AABBVV-3与普通小麦NAU0686杂交、回交的后代中鉴定到一个小麦-簇毛麦2V#6(2D)异代换系11-2V-1,利用11-2V-1在染色体2V和2D之间产生重组,将11-2V-1与高感白粉病普通小麦NAU0686杂交,F_(1)自交。利用GISH/FISH和分子标记对F_(2)世代131个单株进行分子细胞学鉴定,获得了小麦-簇毛麦T2DS·2V#6L易位系11-2V-2、2V#6S端体系11-2V-3和2V#6L端体系11-2V-4新种质。白粉病抗性鉴定发现,所有F_(2)单株苗期均高感小麦白粉菌生理小种E09,但成株期含有2V#6和2V#6L染色体臂的所有单株均表现高抗(IT=1~2),而仅含有2V#6S染色体臂或无外源染色体的单株均高感白粉病(IT=7~9),表明簇毛麦01I139的2V#6L染色体臂上携带成株抗白粉病基因,该基因可能与小麦-簇毛麦T2BS·2V#5L易位系携带的Pm62是等位基因。遗传效应分析表明,T2DS·2V#6L易位染色体在小麦背景传递正常,对主要农艺性状没有显著的负向影响,对穗长、每穗粒数和单株粒重还表现出一定程度的正向效应。因此,本研究创制的成株抗白粉病T2DS·2V#6L易位系11-2V-2为小麦抗白粉病育种提供了新抗源,也为后续簇毛麦2VL上优异基因遗传定位和图位克隆奠定了基础。

荆芥和砂仁挥发油对米象熏蒸活性的协同作用及成分研究

研究20种植物挥发油对米象(Sitophilus oryzae Linnaeus)成虫的熏蒸活性以及混配的协同作用。采用水蒸气蒸馏法提取20种植物挥发油,通过气相-质谱法和化合物消除法,确定熏蒸活性和协同作用最好的2种挥发油的活性成分。结果表明:20种植物挥发油中荆芥挥发油对米象的熏蒸活性最好,LC_(50)为13.27 mg/L。砂仁挥发油表现出最大的协同作用,共毒性因子为107.14。通过化合物消除法评价荆芥挥发油和砂仁挥发油的主要活性成分分别为胡薄荷酮和(-)-4-萜品醇,对米象熏蒸活性的LC_(50)分别为7.34、8.35 mg/L,优于荆芥挥发油的熏蒸活性。胡薄荷酮和(-)-4-萜品醇按照质量比例61.13∶6.13和1∶1的比例混配,LC_(50)分别为5.91、8.29 mg/L,随着胡薄荷酮比例的增大,混剂对米象的熏蒸活性增强。研究证实了胡薄荷酮和(-)-4-萜品醇作为植物源杀虫剂具有防治米象的潜力。

含抗白粉病基因PmV小麦-簇毛麦补偿性插入易位系Dv6-IT2的创制与鉴定

白粉病是中国小麦主要病害之一。抗白粉病基因PmV和Pm21是来自不同簇毛麦种质的同源基因,分别以T6V#4S·6DL和T6V#2S·6AL易位系的形式被育种利用。Pm21基因在中国已得到广泛应用,而作为其后备抗源的PmV基因却极少被利用,主要原因是T6V#4S·6DL易位染色体的外源片段过大,在亲子代传递率低,育种利用困难。创制含PmV的补偿性小片段易位是提高传递率、减少外源基因冗余的重要途径。本研究利用均携带PmV基因的顶端易位系Dv6T25与近着丝粒易位系Dv6T36杂交,根据外源重叠区重组原理,在F_(2)代分离群体中筛选出1个携PmV基因的补偿性中间插入易位系Dv6-IT2。利用簇毛麦参考基因组序列信息进行标记加密分析,结果表明易位片段双侧断裂重接位点分别介于35.1~36.3 Mb和85.1~85.3 Mb,外源易位片段长度约为50 Mb。将Dv6-IT2与大面积推广品种扬麦23杂交构建F_(2)分离群体,标记结果显示,抗感比符合3∶1分离比,说明PmV基因在新型易位中可正常传递。本研究成功创制了Dv6-IT2,将促进PmV基因在小麦抗白粉病育种中的利用。

阳春砂茉莉酸生物合成酶基因鉴定与时空表达分析

为探究阳春砂(Amomum villosum)花器中茉莉酸(JA)生物合成酶基因表达与雌雄蕊生长的关系,本研究利用cDNA末端快速扩增(RACE)技术克隆阳春砂JA生物合成酶基因的全长序列,并对各基因进行生物信息学分析、表达模式测定及各基因表达与阳春砂雌雄蕊生长的相关性分析,以探究阳春砂花器中JA生物合成酶基因表达与雌雄蕊生长的关系。结果表明,阳春砂中JA生物合成酶基因AvLOX、AvAOS、AvAOC、AvOPR3序列全长分别为2733、1798、930、1400 bp,分别包含2577、1485、744、1179 bp的开放阅读框(ORF),其编码蛋白分别含有858、494、247、392个氨基酸残基。4个JA生物合成酶基因的表达随阳春砂雄蕊的发育进程逐渐增加,但在雌蕊中的表达则呈现多样性。在雌蕊中,AvLOX、AvAOS基因在开花后1天表达最高,AvAOC在标记后第9天表达最高,AvOPR3基因在标记后第6天表达最高。除AvAOS基因在小花在标记后第3天以及AvOPR3基因在小花标记后第6天外,4个JA生物合成酶基因在雄蕊中的表达在各发育阶段显著高于同时期在雌蕊中的表达。与对照组相比,在喷施茉莉酸甲酯(MeJA)后,4个JA生物合成酶基因在雄蕊中的表达模式,以及AvLOX和AvOPR3基因在雌蕊中的表达模式没有改变,但AvAOS和AvAOC基因在雌蕊中的表达模式有变化。在对照组和MeJA处理组中,4个JA生物合成酶基因的表达与阳春砂小花花丝和花柱的长度有极显著的相关性。本研究结果为进一步探究JA与阳春砂花器官生长发育的关系奠定了基础。

甲基磺酸乙酯诱变对阳春砂仁出苗的影响

阳春砂仁是我国传统“四大南药”之一。我国春砂仁种质资源单一,遗传基础狭窄,通过甲基磺酸乙酯(EMS)诱变,以期筛选出诱变阳春砂仁种子最合适的EMS浓度和诱变时间,从而为EMS诱变新种质奠定基础。以阳春砂仁种子为试材,采用EMS进行诱变处理,研究了不同EMS浓度、不同处理时间对阳春砂仁种子萌发出苗率和生长势的影响。结果表明:EMS处理体积比浓度为0.2%~0.4%,不同处理时长对出苗数影响较小,但EMS处理浓度为0.8%以上时,随着处理时长的增加,其出苗数大幅度降低,甚至不出苗。因此,EMS对砂仁的生长具有抑制作用。经过研究得出,EMS诱变砂仁的适宜浓度为0.2%~0.4%,适宜时间为4~12 h,EMS处理后55~67 d生长势最快。研究结果可为阳春砂仁化学诱变及新种质资源创制提供参考。

一年生簇毛麦基因资源挖掘与利用研究进展

一年生簇毛麦(Dasypyrum villosum (L.) Candargy)是普通小麦野生近缘种之一,也是近年来应用较广泛的小麦遗传资源。簇毛麦在长期进化过程中,形成了许多可用于小麦遗传改良的重要农艺性状,包括对非生物和生物胁迫的抗性、优良品质等。本文简述了簇毛麦与普通小麦的同源关系、与小麦属杂交亲和性,以及将簇毛麦染色体、片段、基因导入普通小麦的有效方法,综述了簇毛麦对白粉病、纹枯病、条锈病、眼斑病、黄花叶病、全蚀病、胞囊线虫病等病害的抗性基因和对应的染色体,簇毛麦品质基因(如:高赖氨酸含量和多态性贮藏蛋白等)和簇毛麦耐旱、光周期等其他基因。介绍了簇毛麦抗白粉病基因Pm21和PmV在小麦改良和育种中的应用及其巨大价值,展望了簇毛麦后续研究前景和可能存在的问题。本综述对挖掘与利用簇毛麦有益基因、拓宽小麦遗传资源、加快小麦遗传改良进程以及重要基因功能研究均具有参考价值。

春砂仁多糖对高脂饮食小鼠脂代谢和抗氧化作用的影响

探究春砂仁多糖(AVP)对高脂饮食引起的小鼠脂代谢异常和氧化应激的改善作用。试验选取120只6周龄健康的C57BL/6雄性小鼠,随机分为:基础日粮组(CK)、高脂日粮组(HFD)、HFD+AVP低剂量组(HFD+L-AVP)、HFD+AVP中剂量组(HFD+M-AVP)和HFD+AVP高剂量组(HFD+H-AVP),试验共8周。结果表明:日粮中添加AVP,降低了高脂饲喂小鼠血清中AST、ALT、LDL-C、TC和TG的水平,提高了HDL-C水平。提高了小鼠肝脏和肠道中SOD、CAT和GSH-Px的活性,显著降低了肝脏和肠道中MDA水平。降低了高脂饮食小鼠SREBP1和FAS的mRNA表达水平,提高了ATGL的mRNA表达水平,显著抑制小鼠的脂肪生成。AVP的添加显著提高了高脂饮食小鼠肠道中Nrf2、HO-1 mRNA和蛋白的表达水平,也提高其肠道抗氧化能力。综上所述,春砂仁多糖可缓解高脂饮食导致的小鼠氧化应激和脂代谢异常。