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1.
一种高效提取猕猴桃DNA和RNA的方法 总被引:1,自引:0,他引:1
在总核酸提取方法(PS法)的基础上,经多次实践改进,得出一种以高盐低pH的HAc-NaAc缓冲体系提取总核酸的简便方法,可以从富含多糖、多酚时猕猴桃叶片和花蕾中提取同时含有DNA和RNA的总核酸.所得的总核酸在LiCl溶液中选择性沉淀RNA,从而有效地分离出DNA和RNA样品.紫外分光光度法和琼脂糖凝胶电泳分析表明,所提取的DNA和RNA具有较高的纯度和完整性.通过样品DNA的PCR和样品RNA的RT-PCR,认为所提取的DNA样品和RNA样品能够满足分子生物学试验的基本要求. 相似文献
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提取北方土壤真菌DNA的一种方法 总被引:6,自引:1,他引:6
由于土壤理化性质的复杂性和真菌细胞壁结构的特殊性,从土壤样品中提取真菌基因组DNA比较困难.中国北方土壤与其它地区土壤相比有其自身的特点,因此,有必要优化一种适合于北方土壤真菌DNA提取的方法.本实验向灭菌黑土中分别投加12种在系统分类上差别较大的真菌,以传统土壤总DNA提取方法及纯菌DNA提取方法为基础,分别与蜗牛酶,纤维素酶进行组合、优化,得到7种不同的土壤真菌基因组DNA提取方法.利用真菌28S rDNA通用引物U1/U2-GC PCR-DGGE分析方法分别考察了7种不同方法所提取土壤真菌基因组DNA的多样性和代表性.结果表明:1)液氮研磨,纤维素酶、蜗牛酶和溶菌酶(浓度分别为6 、3和1 mg·ml-1)37 ℃作用60 min,2% SDS于65 ℃裂解30 min;2)-65 ℃~65 ℃冻融3次,纤维素酶、蜗牛酶和溶菌酶(浓度分别为6、3和1 mg·ml-1)37 ℃作用180 min,2%SDS于65 ℃裂解30 min的组合具有较好的提取效果.利用后一种方法分别对3种理化性质差异较大的中国北方自然土壤样品真菌DNA进行提取并分析,表明所提取土壤基因组DNA真菌特异性PCR-DGGE图谱条带丰富,该方法可用于多种北方土壤真菌多样性研究. 相似文献
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为研究玻璃粉在植物核酸提取中的应用,比较了玻璃粉颗粒大小、离液盐种类及浓度、pH等条件对玻璃粉吸附核酸的影响,得出玻璃粉吸附核酸的各种最佳条件。结果表明,普通玻璃粉吸附核酸能力强于硅胶和硅藻土,玻璃粉颗粒的直径以83 μm为佳,pH 4.0时吸附效果达到最大。提取DNA时,NaCl浓度应大于3 mol/L,而提取RNA时,异硫氰酸胍大于2 mol/L就能取得很好的效果,此外,在玻璃粉吸附RNA前,需要加入50%以上的无水乙醇才能更好地吸附。利用玻璃粉制作简易纯化柱,可用于植物组织核酸提取纯化,所提取的核酸纯度高、完整性好,可用于酶切、杂交和PCR等实验。与传统方法相比,采用玻璃粉简易离心柱提取植物核酸,效果好、环保、快速、经济。 相似文献
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一种简单有效且适于土壤微生物多样性分析的DNA提取方法 总被引:1,自引:0,他引:1
参照Zhou[11]的方法进行了改进,获得了一种简单、有效的DNA提取方法.此方法操作简单、从大量样品改为小量样品的提取,利用高浓度的PEG沉淀,不作回收纯化,所提DNA片段较大,在23 kb以上,每克土的DNA提取量从3.74~15.28 μg,OD260/OD230比值在0.89~1.21范围内,用真菌和细菌核糖体特异性引物进行PCR扩增,均获得较好的结果,DGGE图谱显示丰富性较高,可用于细菌多样性和真菌多样性的分析.此方法能够从4种不同性质土壤中提取出DNA,但提取盐渍土壤和碱性土壤的效果更好一些,为土壤微生物群落结构的多样性分析奠定良好的基础. 相似文献
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《中国真菌学杂志》2017,(6)
目的探索一种新的真菌DNA提取方法,该方法操作简便,提取效率高,耗时较短,应用范围广。方法将酶消化和Chelex-100提取DNA方法联合起来,配制一种新的DNA提取试剂。通过真菌ITS4和ITS5通用引物进行PCR,对新的DNA提取试剂同市售的Takara lysis buffer在DNA提取方面的效能进行评价,同时对DNA浓度和纯度进行测定。结果实验所用34株真菌,自配试剂成功提取出32株真菌DNA;Takara lysis buffer成功提出27株真菌DNA。对于两种方法都没能提出DNA的2株真菌,经液氮研磨破壁后,自制试剂均成功提取;而Takara lysis buffer仅成功提取DNA 1株。结论同市售试剂Takara lysis buffer比较,自配DNA提取液提取DNA质量相对较高,可用于临床常见感染真菌的PCR诊断。对于某些真菌,提取DNA之前先进行菌体破壁是必要的。 相似文献
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副溶血弧菌(Vibrio parahaemolyticus)是一种革兰氏阴性嗜盐菌.提取高质量的RNA是研究副溶血弧毒力基因表达与其致病性和环境适应性的基础.本研究分别用SDS法、Trizol法,PureLinkTM RNA Mini Kit和Uniq-10柱式Trizol总RNA抽提试剂盒提取六株副溶血弧菌的总RNA,用普通琼脂糖凝胶电泳和核酸测定仪检测RNA的质量,并用RT-PCR法对四种提取方法进行比较.研究结果表明,Trizol法和PureLinkTM RNA Mini Kit简单快捷,提取的总RNA完整性好,纯度高,可用于后续实验的分子生物学研究;而Trizol法更适用于普通实验室细菌RNA的提取. 相似文献
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为筛选和建立风沙土中总DNA的提取和纯化方法,选取了5种直接提取法、1种间接提取法和2种纯化法分别对风沙土中总DNA进行了提取和纯化,并对其质量和产量进行了比较.结果表明:6种方法均可从风沙土中提取到大小为23 kb左右的总DNA,其中改进后的高盐提取法(用40%聚乙二醇8000和4 mol·L-1 NaCl沉淀DNA)效果最好,纯化后总DNA的纯度最高,可进行16S rDNA的PCR扩增,且产量仅稍低于试剂盒提取法;电泳加柱回收纯化法的纯化效果较好,经该方法纯化后的总DNA大部分可进行PCR扩增,可满足后续分子操作对DNA纯度的要求. 相似文献
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目的 为快速地提取到质量较好的黑翅土白蚁基因组DNA进行白蚁种群多样性的研究,对基因组DNA提取方法进行了比较与改进.方法 先初步采取CTAB法与蛋白酶K法对黑翅土白蚁基因组DNA的提取方法进行比较,再利用正交设计法对蛋白酶K法中裂解液、蛋白酶、RNA酶及作用时间4个因素进行优化.结果 蛋白酶K法获得的基因组DNA的质量与产量稍优于CTAB法;较佳的提取步骤组合为:裂解液150 μL,蛋白酶K 6μL,作用时间1h,RNA酶可不添加.结论 采用优化后的方法获得的基因组DNA为模板进行PCR扩增,得到了清晰、稳定的扩增谱带,完全可用于相关后续实验. 相似文献
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Pushkar Shejwalkar 《Inorganica chimica acta》2011,366(1):209-218
New sulfur derivatives of phosphoramidite ligands were synthesized and the impact of the sulfur unit on the spectroscopic properties of their rhodium and iridium complexes was investigated. The new ligands Bn2NPSCH2CH2Sa(P-Sa) (Bn = benzyl, 4), Bn2NPSCHCHSa(CH2)3CaH2(P-Sa)(Ca-Sa) (6) and Bn2NP(4-XC6H4OMe)2 (X = S, 7a; X = O, 7b) were converted to the rhodium and iridium complexes trans-[Rh(CO)Cl(L)2] (L = 4, 6, 7), [RhCl(COD)(L)] (L = 4, 6, 7), [IrCl(COD)(7a)] and [IrCl2Cp∗(6)]. For comparison, some phosphoramidite complexes of these formulations also were synthesized. The new metal complexes were spectroscopically analyzed. For the carbonyl complexes, the νCO IR stretching frequencies were lower than for the corresponding phosphite and phosphoramidite ligands. The 1JPRh coupling constants for the rhodium complexes with the new ligands were also smaller than for the respective phosphoramidite and phosphite complexes. Finally, the 1JPSe coupling constants of the selenides of the new ligands were lower than those of the phosphoramidite ligands but higher than for PPh3. The spectroscopic data reveal that the new thio ligands 4, 6 and 7a are more electron donating than phosphites and phosphoramidites but less electron donating than PPh3. 相似文献
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Astrocytes transport the monocarboxylate acetate, but synaptosomes do not. The reason for this is unknown, because both preparations express monocarboxylate transporters (MCT). The transport and metabolism of lactate, another monocarboxylate, was examined in these two preparations, and the results were compared to those for acetate. Lactate transport is more rapid in astrocytes than in synaptosomes, but of lower affinity (Kms of 17 and 4 mM, respectively). Lactate (0.2 mM) is metabolized to CO2 more rapidly in synaptosomes than in astrocytes (rates of 0.37 and 0.07 nmol x mg protein(-1) x min(-1), respectively). The reason for this is unclear, but cellular differences in lactate dehydrogenase isotype expression may be involved. Acetate is metabolized to CO2 more rapidly in astrocytes than in synaptosomes (rates of 0.43 and 0.02 nmol x mg protein(-1) x min(-1), respectively). This is likely due to cellular differences in the expression of monocarboxylate transporter subtypes. 相似文献
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Hand JW 《Progress in biophysics and molecular biology》2007,93(1-3):192-194
The first and second sessions of the Workshop focussed on the basics of ultrasound and infrasound, their applications in both industry and medicine, and metrology and protection standards for ultrasound applications. 相似文献
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Gonzalez-Reimers E Rodriguez-Moreno F Martinez-Riera A Mas-Pascual A Delgado-Ureta E Galindo-Martin L Arnay-de la Rosa M Santolaria-Fernandez F 《Biological trace element research》1999,68(1):41-49
To elucidate accumulation of minerals in human iliac arteries with aging, the content of minerals was analyzed by inductively
coupled plasma atomic emission spectrometry. Bilateral common, internal, and external iliac arteries of 16 men and 8 women,
ranging ages from 65 to 93 yr, were examined. It was found that an extremely high accumulation of calcium and phosphorus occurred
in the common iliac artery at old age, being higher than that of the internal and external iliac arteries. It should be noted
that the accumulation of calcium and phosphorus is the highest in the common iliac artery among the human arteries examined
to date.
Regarding sexual differences, the content of calcium and phosphorus in the common and internal iliac arteries was higher in
women than in men, whereas their content in the external iliac artery was lower in women than in men. 相似文献
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J. Morley M.A. Bray R.W. Jones D.H. Nugteren D.A. van Dorp 《Prostaglandins & other lipid mediators》1979,17(5):729-736
The ability of partially purified human and guinea-pig haematogenous cell populations, when cultured in vitro, to metabolise arachidonic acid (AA) has been studied. Supernatants from 24 hour cell culture have been subjected to analysis for products of AA metabolism by gas chromatography with electron-capture detection.The cell types studied were human peripheral blood monocytes (both glass adherent and non-adherent), neutrophils, eosinophils and leukemic leucocytes; thoracic duct lymphocytes and lung alveolar macrophages. From the guinea-pig, induced and non-induced macrophage or neutrophil enriched peritoneal exudate populations, lymph node cells, peritoneal eosinophils and peripheral blood platelets were examined. Supernatants were assayed for the presence of PGE2, PGD2, PGF2α, TXB2 and 6-keto-PGF1α. In all types studied PGE2 and TXB2 were the major products formed. The identification of PGE2 and TXB2 was confirmed by GC/MS with multiple ion monitoring.The results have been compared with other reports and their possible significance discussed in relation to the proposed role of prostaglandins as mediators and modulators in immunopathology. 相似文献
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Allergic asthma can be precipitated by many factors. For the atopic person, fungus, pollen, dust mites, cockroach antigens, and diesel exhaust are all agents that may trigger an allergic attack. Cytokines and chemokines are integral mediators of fungal asthma. From the earliest time points, they recruit and activate the cells required for the clearance of fungus as well as being critical factors involved in the immunopathology of this disease. In the final analysis, it is clear that these mediators can act to the benefit or the detriment of the host. 相似文献
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Sandra M. Vaz 《Archives of biochemistry and biophysics》2009,484(2):127-431
In spite of the many studies on protein modifications by reactive species, knowledge about the products resulting from the oxidation of protein-aromatic residues, including protein-derived radicals and their stable products, remains limited. Here, we compared the oxidative modifications promoted by peroxynitrite and myeloperoxidase/hydrogen peroxide/nitrite in two model proteins, ribonuclease (6Tyr) and lysozyme (3Tyr/6Trp). The formation of protein-derived radicals and products was higher at pH 5.4 and 7.4 for myeloperoxidase and peroxynitrite, respectively. The main product was 3-nitro-Tyr for both proteins and oxidants. Lysozyme rendered similar yields of nitro-Trp, particularly when oxidized by peroxynitrite. Hydroxylated and dimerized products of Trp and Tyr were also produced, but in lower yields. Localization of the main modified residues indicates that peroxynitrite decomposes to radicals within the proteins behaving less specifically than myeloperoxidase. Nitrogen dioxide is emphasized as an important protein modifier. 相似文献