沙冬青cDNA-AFLP银染和条带回收方法分析

通过比较而获得沙冬青cDNA-AFLP银染和条带回收的最佳方法.银染时采取Bassam法和Sanguinetti法,凝胶条带回收时利用直接回收法、试剂盒法、LiCl高盐法和聚丙烯酰胺凝胶高效回收法.比较不同方法对于开展聚丙烯酰胺凝胶电泳的影响.采用Bassam银染法对获得的扩增产物进行染色后无法得到差异显示条带,而利用Sanguinetti银染法显色后可观察到比较明显的差异表达条带;以直接回收法、PAGE试剂盒法、LiCl高盐法对差异条带进行回收后,二次PCR无法得到扩增产物,采用聚丙烯酰胺凝胶高效回收法后则可获得比较清晰的二次扩增产物.Sanguinetti银染法和聚丙烯酰胺凝胶高效回收法是应用于本研究的最佳方法.     

一种高效、快速从凝胶中回收DNA的方法

目的:介绍了一种从普通琼脂糖电泳中回收DNA的简便、快捷、高效且廉价的方法.方法:利用0.5 mL离心管、1.5mL离心管、尼龙膜做成的一个小装置.把含有DNA的凝胶放在膜上,离心,收集从管底流出来的液体,用乙醇沉淀DNA.结果:最终回收率为60%左右,回收率大约为市售试剂盒的90%,接近市售DNA回收试剂盒.结论:该方法操作简单,回收率高,无其他试剂污染.     

Beacon公司微囊藻毒素检测试剂盒的性能评价

对一种进口微囊藻毒素ELISA试剂盒进行应用性能评价。用该试剂盒进行精密度实验,标准品添加回收实验,交叉反应实验以及样品检测比较实验。试剂盒的分析内检测精密度较高,分析间检测精密度偏低,加标回收率在73.5-97.8%之间,试剂盒抗体与MC-LR的交叉反应率很高,但与MC-RR、MC-LW、MC-LF等微囊藻毒素异构体交叉反应率偏低,在水样的测定中,试剂盒检测结果与本实验室方法检测结果基本一致。该试剂盒基本能够满足对水体中MC-LR的定性和定量检测要求。     

四种土壤微生物总DNA的纯化方法的比较

比较了4种从土壤中直接抽提的微生物总DNA的纯化方法,实验结果表明1 %的琼脂糖凝胶电泳纯化方法及葡聚糖凝胶G 2 0 0离心层析纯化方法均不能完全纯化从土壤中抽提的微生物总DNA。若将直接抽提的总DNA先经葡聚糖凝胶G 2 0 0离心层析纯化,再用1 %的琼脂糖凝胶电泳纯化,则能取得较好的纯化效果。含2 %PVP的1 %琼脂糖凝胶电泳纯化,用DNA凝胶回收试剂盒回收后没有得到纯化后的土壤微生物总DNA。     

利用机油提高琼脂糖凝胶中DNA片段的回收率

目的:发展一种简单快速经济的回收琼脂糖凝胶中DNA的方法.方法:将切的琼脂糖凝胶胶块放入嵌套Eppendorf管中捣碎,加入50μL机油,室温下12 000r/min离心5min,取大Eppendorf管中收集的液体跑胶检验,凝皎上包括代表100bp～2000bp不同大小DNA分子回收率的分子量标准DL2000.结果:DNA回收率可由加机油前的约35%提高为加机油后的45%-90%,回收效率的波动主要取决于DNA片段的大小、切下的含有DNA的胶块的大小及操作者的熟练程度.结论:该方法快速、简便、经济,具有良好的重复性与特异性,比许多国产的试剂盒更可靠.     

简易电泳洗脱蛋白方法

制备电泳的连续洗脱与切下目的蛋白谱带洗脱是从聚丙烯酰胺凝胶回收蛋白质常用的两种方法。切带回收蛋白可采用浸泡提取,化学解聚凝胶及电泳洗脱多种途径。相对比较,电泳洗脱蛋白回收率较前二者高些,因此人们多采用此法。但是,目前广泛使用     

一种从琼脂糖凝胶中同步回收DNA和琼脂糖的方法

介绍一种从琼脂糖凝胶同步回收DNA和琼脂糖的方法。利用0.25 mol/L异硫氰酸胍溶液(p H 8.0)溶解含有目的 DNA片段的的凝胶条,胶条溶解后,静置冰上10 min再加入预冷的异丙醇,琼脂糖呈颗粒状析出,通过离心即可初步分离DNA和琼脂糖。上清液用异丙醇沉淀回收DNA片段,利用50%PEG溶液沉淀琼脂糖。分别对0.2 kb、1 kb和10 kb长度的DNA片段进行回收,回收率分别为19.44%、36.40%、13.49%,回收的DNA纯度高,电泳条带清晰。琼脂糖均回收率为62.52%,回收琼脂糖脱水后的状态为白色颗粒。该方法切实可行,回收成本低廉,回收的DNA和琼脂糖可用于后续实验。     

蛇毒纤溶酶Fibrolase的凝胶过滤色谱复性研究

目的：研究蛇毒纤溶酶Fibrolase的最佳复性方法。方法：使用透析和凝胶色谱法对Fibrolase复性进行研究,比较2种复性方法的相对复性率及蛋白质回收率。结果：2种复性方法的复性率分别为20％、25％,蛋白质回收率分别为16％、5％。结论：凝胶过滤色谱复性优于透析法复性,凝胶过滤色谱复性使蛋白质的复性与纯化同步进行,简化了操作程序,提高了产品回收率。     

简便实用的琼脂糖凝胶回收DNA片段方法

介绍一种简便实用的DNA片段回收方法,与以前所报道的DEAE-纤维素膜电泳法、透析袋电洗脱法、低融点琼脂糖凝胶法、凝胶冻融法等相比,所需器材简单、操作简便、回收率高、成本低。回收的DNA片段在进一步克隆和测序中表现出较好的效果,是一种适合于科研和教学的实验方法。     

CTAB结合DNA凝胶回收试剂盒提取食用菌DNA

目的:为探索从食用菌子实体中快速分离和纯化DNA的方法.方法:以三种栽培的食用菌为材料,采用CTAB结合DNA凝胶回收试剂盒进行基因组DNA的分离和纯化.结果:与对照相比,结合法提取DNA样品的电泳条带更规则、清晰,并且DNA的酶切效率显著高于对照.结论:该结合法是一种快捷、高效提取纯化食用菌子实体DNA的方法.     

Decrease in Cryptosporidium parvum oocyst infectivity in vitro by using the membrane filter dissolution method for recovering oocysts from water samples.

Exposure of Cryptosporidium parvum oocysts to solutions used for cellulose acetate membrane (CAM) dissolution filtration reduced their infectivity in HCT-8 cells. Ethanol (95% [vol/vol] and 70% [vol/vol]) alone and short exposure times to acetone decreased infectivity. These findings contrast with similar experiments using excystation assays and infectivity in mice.     

Ischemia-reperfusion injury--antiarrhythmic effect of melatonin associated with reduced recovering of contractility

The effect of melatonin on reperfusion arrhythmias and postischemic contractile dysfunction was studied in the isolated rat heart. 25 min global ischemia was induced and followed by 30 min of reperfusion. Melatonin (10 micromol/l) was present in the perfusion solution during the whole experiment. Experiment revealed protective effect of melatonin on reperfusion-induced arrhythmias--arrhythmia score was significantly lower as well as the total time of arrhythmias duration was significantly shorter in melatonin group than in controls. On the other hand, post-ischemic recovering of contractility was significantly reduced in melatonin group.     

31P-MRS study of bio-energy recovering phenomenon

The ATP and creatine phosphate (PCr) contents in isolated guinea-pig hearts were determined by 31P-MRS measurement at 80.75 MHz using the Langendorff technique. Reperfusion of post-ischemic hearts with adenosine for 180 minutes increased ATP to 117.4% and decreased PCr to 59.8% of the preischemic value. Reperfusion without adenosine did not increase ATP and did not decrease PCr. The depressed cardiac function due to ischemia was remarkably improved in post-ischemic hearts by the increase in ATP due to adenosine. We found that the loss of ATP due to ischemia is not necessarily proportional to the extent of myocardial ischemic injury.     

Multi-stress resilience in plants recovering from submergence

Submergence limits plants' access to oxygen and light, causing massive changes in metabolism; after submergence, plants experience additional stresses, including reoxygenation, dehydration, photoinhibition and accelerated senescence. Plant responses to waterlogging and partial or complete submergence have been well studied, but our understanding of plant responses during post-submergence recovery remains limited. During post-submergence recovery, whether a plant can repair the damage caused by submergence and reoxygenation and re-activate key processes to continue to grow, determines whether the plant survives. Here, we summarize the challenges plants face when recovering from submergence, primarily focusing on studies of Arabidopsis thaliana and rice (Oryza sativa). We also highlight recent progress in elucidating the interplay among various regulatory pathways, compare post-hypoxia reoxygenation between plants and animals and provide new perspectives for future studies.     

Numerical experiments with a new differentiation filter

A dynamic programming filter which provides estimates of the first and second derivative of empirical displacement data is investigated numerically. This filter uses a weighted least squares criteria in estimating the derivatives. The filter equations are presented together with several numerical examples. These examples are taken from references that proposed other techniques.     

Upflow biological filtration with floating filter media

An aerobic biological filter with floating filter media was tested using domestic wastewater to determine the optimum operating and backwashing parameters in this study. This system was designed for a small-scale joint treatment plant and 4 mm diameter polystyrene foam pellets were used as the filter media. As the backwash is the most important point for using a floating biofilter, “air shot” system, a turbulent-flow backwashing utilising the air reverse syphon and the buoyancy of the filter media, was developed for this purpose. Through the test, the optimum position of the diffuser tube and the structure of the backwashing device were determined. It was confirmed that the system could achieve effluent biological oxygen demand (BOD) of under 10 mg/l, and a nitrification rate of over 86% at a BOD loading of 0.7 kg/m³ per day and “total-nitrogen (T-N) loading” of 0.16 kg/m³ per day. The backwashing frequency should be as many times as possible per cycle to remove the sloughed sludge thoroughly in order to maintain good effluent quality after backwashing.     

Methods for recovering poliovirus and rotavirus from oysters

Polioviruses and rotaviruses are potential indicators of sewage pollution of water and shellfish. Several methods for detecting these viruses in oysters were assessed. Elution-precipitation involving Catfloc for clarification and skim milk for subsequent flocculation resulted in the recovery of an average of 79% of poliovirus type 1 and 37% of rotavirus SA-11 from oyster homogenates inoculated with low numbers of these viruses. Adsorption-elution-precipitation did not improve the recovery of poliovirus and was detrimental to the recovery of rotavirus. Ultrafiltration or ultracentrifugation resulted in improved recovery of rotavirus but also in higher toxicity of oyster extracts to cell cultures. We recommend the use of the described elution-precipitation method for detecting viral pollutants in sample of oysters.