适合于基因组测序的红麻高纯度线粒体DNA提取

采用红麻黄化苗为试材,结合密度梯度离心和差速离心的方法提取线粒体DNA(mtDNA)以满足测序要求.结果表明,蔗糖密度梯度离心法比Percoll密度梯度离心法更适合于红麻线粒体的分离.分离后的线粒体经DNaseI消化核DNA,并采用Jannus绿检测线粒体的完整性,SDS和蛋白酶K裂解线粒体,并用酚/氯仿抽提除去蛋白,...     

一种家蚕后部丝腺高纯度线粒体提取方法

为了从家蚕后部丝腺中提取高纯度的线粒体,采用差速离心和非连续Percoll密度梯度离心方法,分离和纯化家蚕后部丝腺线粒体,并通过电镜观察和蛋白免疫印迹验证提取效果。结果表明,采用该方法分离的线粒体杂质少、纯度高,并且线粒体内外膜结构完整。该研究建立了一种提取家蚕后部丝腺线粒体的方法,为进一步研究线粒体在家蚕后部丝腺中的功能奠定了基础。     

超滤离心在双水相萃取技术中的应用

双水相萃取技术作为新型分离技术日益受到重视,但由于萃取物与成相物质分离难等限制其应用。文章论述双水相萃取技术与超滤离心技术的特点、原理及其应用,分析两种分离技术结合使用可能会出现的问题,重点阐述超滤离心在双水相萃取技术中分离和回收产物时的工艺方法。     

蔗糖离心法分离土壤线虫的最佳条件筛选

采用单因素试验和二因素组合试验,对蔗糖离心法分离线虫的效果进行研究。分别对在蔗糖液中的离心时间、离心转速和蔗糖的浓度进行了单因素试验,并对效果较好的离心时间和离心转速进行了二因素组合设计试验。试验结果表明:离心时间和离心速度对线虫的分离有影响,但蔗糖浓度对分离结果影响不显著。经蔗糖离心30s后,所获得的线虫数量最多;离心2000r.min-1(离心力447×g)时,效果显著高于其他4个离心转速处理;在离心时间和离心转速的二因素组合试验中,以30s、2000r.min-1的分离效果为最优。     

牛精子不同获能方法对体外受精效果的影响（英文）

为了提高牛体外受精率,探讨牛冷冻精液不同获能方法对体外受精效果的影响,对牛冷冻解冻精液进行了两次离心获能法与一次离心加上游法获能对体外受精效果影响试验。结果表明,两次离心组受精率卵裂率(75.6±4.5)%与一次离心加上游法组受精卵裂率(76.4±1.9)%无显著差异(P0.05);二次离心组囊胚率(35.7±4.1)%与一次离心加上游法组(36.3±2.7)%差异不显著(P0.05)。而两次离心组却可以大大节省了体外获能时间。     

牛精子不同获能方法对体外受精效果的影响

为了提高牛体外受精率,探讨牛冷冻精液不同获能方法对体外受精效果的影响,对牛冷冻解冻精液进行了两次离心获能法与一次离心加上游法获能对体外受精效果影响试验.结果表明,两次离心组受精率卵裂率(75.6-±4.5)％与一次离心加上游法组受精卵裂率(76.4±1.9)％无显著差异(P＞0.05);二次离心组囊胚率(35.7±4.1)％与一次离心加上游法组(36.3±2.7)％差异不显著(P＞0.05).而两次离心组却可以大大节省了体外获能时间.     

鹿血总蛋白分离工艺参数研究

通过影响鹿血血浆和血细胞分离效果的单因素试验,确定切实可行的分离工艺。方法是以血浆得率、血浆中总蛋白浓度和血红蛋白浓度为指标,研究离心转速和离心时间对分离效果的影响。结果是在离心转速3500r/min,离心时长15min时,血浆得率高,血浆中总蛋白质量浓度高,血红蛋白浓度低,离心分离效果最好。并且该分离参数下分离成本低,分离效果好。     

茭白线粒体蛋白双向电泳体系建立

[目的]建立适合于茭白线粒体蛋白质组分析的双向电泳技术体系,为进一步开展茭白线粒体差异蛋白质组学研究提供参考.[方法]以茭白为试验材料,比较不同线粒体提取纯化方法、IPG胶条pH范围及蛋白上样量等条件对双向电泳图谱的影响.[结果]差速离心(3000~14000×g)联合Percoll不连续密度梯度离心(20%∶24%∶40%=2∶4∶2)对茭白线粒体的提取纯化效果优于仅采用差速离心.采用改良酚抽法提取茭白线粒体蛋白,17 cm、pH 3~10的IPG胶条,1.0 mg上样量,12% SDS-PAGE进行双向电泳,0.12%考马斯亮蓝G-250胶体考染法染色,茭白线粒体蛋白主要分布在pH 4~8范围内,pH 3~4和pH 8~10范围内蛋白点较少;进一步选用pH 4~7和pH 5~8的IPG胶条对茭白线粒体蛋白进行双向电泳,分离效果均明显提高,且pH 4~7范围内IPG胶条的双向电泳图谱清晰,蛋白点分辨率较高.分别采用0.8、1.0和1.2 mg 3个不同的上样量进行双向电泳,结果表明上样量为1.0 mg时,电泳图谱质量最好.[结论]通过差速离心联合Percoll不连续密度梯度离心提取纯化茭白线粒体,并控制好双向电泳体系的pH范围、上样量等因素,可获得蛋白质点清晰、分辨率高、重复性好的双向电泳图谱.     

小白菜线粒体DNA提取体系的建立

为了建立小白菜线粒体DNA(mtDNA)提取体系,以小白菜黄化苗为材料,研究了不同提取方法、材料选择等因素对线粒体DNA提取的影响。结果表明,经冰浴研磨小白菜黄化苗,通过简易的密度梯度离心技术快速抽提大量高纯度的线粒体,蛋白酶K裂解线粒体及有机溶剂抽提去除蛋白质,可获得纯度较高的mtDNA,能用于RAPD分析。     

热水解剩余污泥离心除沙试验研究

[目的]研究离心机转速的差异对热水解剩余泥沙分离效果的影响。[方法]两种不同含固率(TS)的热水解污泥(TS分别为13.22和29.42)分别在转速12 000、8 000、4 000、3 000、1 500 r/min条件下离心20 min,取上清液、污泥层和沙粒层样品测定含固率TS和挥发性固体VS。[结果]随着转速的提高,热解泥的泥水分离效果和泥沙分离效果逐渐变好。在离心转速达到8 000 r/min时,两种含固率的热解泥无机沙粒层的TS分别达到77.49%和78.22%,而相应的有机质含量(VS)则分别降为4.05%和5.13%。[结论]研究表明经8 000 r/min离心除沙即可取得较好的效果,因此经济合理的离心条件为转速8 000 r/min,持续时间20 min。     

Southern corn leaf blight: susceptible and resistant mitochondria

Mitochondria isolated from etiolated shoots of blight-susceptible and blight-resistant corn plants were subjected, in various respiratory states, to the pathotoxin released by Helminthosporium maydis (race T). The addition of the pathotoxin to susceptible mitochondria caused respiratory rate and oxidative phosphorylation changes. The addition of pathotoxin to susceptible mitochondria suspended in a potassium chloride reaction medium induced an immediate and irreversible swelling regardless of the respiratory state of the mitochondria. This membrane swelling correlates with the observed respiratory and coupling effects of the pathotoxin. In all instances, mitochondria isolated from blightresistant corn failed to exhibit any of the above responses to the pathotoxin.     

Mitochondrial autonomy: incorporation of monosaccharides into glycoprotein by isolated mitochondria

Isolated intact mitochondria selectively incorporate monosaccharides from nucleotide diphosphate monosaccharides into protein. Fucose, mannose, glucose, and galactose were incorporated by the mitochondria into glycoprotein; xylose was not. Structural integrity of the mitochondria was not necessary for the incorporation of monosaccharide into glycoprotein; mitochondria broken by homogenization also incorporated monosaccharide. The monosaccharides incorporated into glycoprotein were localized in the inner mitochondrial membranes, the same membranes which contain the protein into which leucine is incorporated by the isolated mitochondria.     

Liver mitochondria from manganese-deficient and pallid mice: function and ultrastructure

Oxidative phosphorylation was studied in isolated liver mitochondria from manganese-deficient mice and in those from a mutant strain, pallid. In mitochondria from manganese-deficient mice, ratios of adenosine triphosphate formed to oxygen consumed were normal, but oxygen uptake was reduced. Electron microscopy of these mitochondria revealed ultrastructural abnormalities including elongation and reorientation of cristae. No biochemical or structural abnormalities were found in mitochondria from pallid mice.     

Beta-hydroxybutyrate dehydrogenase: lack in ruminant liver mitochondria

The enizyme D(-)-beta-hydroxybutyrate dehydrogenase has been reported to occur in all mitochondria isolated from mammalian tissues and is especially high in liver. By contrast only very low concentration of the enzyme is detectable in mitachondria from bovine and sheep liver. The amount of the enzyme in mitochondria from other ruminant tissues such as kidney and heart are comparable with those from mammalian sources. The verv low activity of beta-hydroxybutyrate dehydrogenase is not referable to the iniability of substrates to penetrate milochondria or to decreased electron transfer activity but rather to a lack of the enzyne in ruminant liver mitochondria.     

THYROID HORMONES: CONTROL OF TERMINAL OXIDATION

Triiodothyronine administered to thyroidectomized rats preferentially increased the total capacity of the electron transport phosphorylation system when isolated mitochondria from the liver were tested with different substrates, but it caused little increase in the activity of the slowest dehydrogenases and no uncoupling. The increased activity appeared to be partly due to direct activation of some component of the electron transport system, although triiodothyronine injection also stimulated the incorporation of amino acids by isolated mitochondria.     

Dimethyl sulfoxide protects tightly coupled mitochondria from freezing damage

Dimethyl sulfoxide prevented loss of respiratory control and decrease in efficiency of oxidative phosphorylation when plant mitochondria were stored in liquid nitrogen. Respiration was severely inhibited and was not stimulated by adenosine diphosphate when mitochondria were frozen in liquid nitrogen without dimethyl sulfoxide. Thus, isolated mitochondria provide a model system for the study of the effects of freezing on biological membranes and of the prevention, by dimethyl sulfoxide, of freezing damage.     

A 13-kilodalton maize mitochondrial protein in E. coli confers sensitivity to Bipolaris maydis toxin

The Texas male-sterile cytoplasm (cms-T) of maize carries the cytoplasmically inherited trait of male sterility. Mitochondria isolated from cms-T maize are specifically sensitive to a toxin (BmT-toxin) produced by the fungal pathogen Bipolaris maydis, race T, and the carbamate insecticide methomyl. A mitochondrial gene unique to cms-T maize, which produces a 13-kilodalton polypeptide associated with cytoplasmic male sterility, was expressed in Escherichia coli. After addition of BmT-toxin or methomyl, inhibition of whole cell respiration and swelling of spheroplasts were observed in Escherichia coli cultures producing the novel mitochondrial protein; these effects are similar to those observed with isolated cms-T mitochondria. The amino-terminal region of the 13-kilodalton polypeptide appears to be essential for proper interaction with the BmT-toxin and methomyl. These results implicate the 13-kilodalton polypeptide in conferring toxin sensitivity to mitochondria of cms-T maize.     

Mitochondrial membrane potentials measured with microelectrodes: probable ionic basis

The membrane potentials of isolated Drosophila mitochondria have been measured under a variety of conditions by microelectrodes driven by a piezoelectric device. The results support the interpretation that the potentials are the result of a distribution of ions imposed by a Donnan effect.     

家兔肝脏线粒体微量量热研究——腐植酸对线粒体代谢热谱的影响

利用ＬＫＢ－２２７７生物活性检测系统测定了家兔肝脏线粒体正常代谢的谱及腐植酸对其代谢热谱的影响，发现线粒体从细胞分离出来来后具有一定活性，可存活较长一段时间，其代谢热谱与细菌热谱相似，可分为抑制期，恢复期，稳定期和衰蝎期，腐植酸对线粒体热谱有较大的影响可使产热提前，产热最高峰增大，总产热量增加，探讨了腐植酸影响线粒体产热的原因。     

Respiratory control: loss in mitochondria from diseased plants

Mitochondria from healthy oat seedlings oxidized succinate with good respiratory control and high ratios of adenosine diphosphate to oxygen. After treatment with victorin, the pathotoxin responsible for symptoms of Victoria blight of oats, susceptible seedlings yielded mitochondria with little respiratory control and lower ratios of adenosine diphosphate to oxygen. No such effects were obtained with victorin-treated resistant seedlings or when victorin was added directly to mitochondria from healthy susceptible or resistant plants. These data indicate that victorin-induced disease results in a reduction in efficiency of the energy-generating system of isolated mitochondria.