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全能型植物RNA提取试剂盒 (DNase I)
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| 货号 (SKU) | 包装规格 | 是否现货 | 价格 | 数量 |
|---|---|---|---|---|
| O665690-50T |
50T |
期货  |
|
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基本描述
| 英文名称 | OminiPlant RNA Kit (Dnase I) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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| 储存温度 | -20°C储存 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| 运输条件 | 超低温冰袋运输 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| 产品介绍 |
产品简介 本试剂盒适用于从多种植物中提取RNA，即便是从多糖多酚含量丰富的植物中也能成功提取高质量的RNA，如水稻叶片、小麦叶片、玉米叶片、烟草叶、松针、银杏叶、杨树叶、石榴叶、冬青叶、苹果、桃、梨、西红柿、樱桃、杏、香蕉、葡萄、枇杷、桂圆果皮、桂圆果肉、荔枝果皮、荔枝果肉、大豆、花生、玉米、马铃薯块茎、月季花瓣、石榴花瓣，香菇、平菇等样本。独特的裂解液配方，可使细胞中的RNA酶迅速灭活，有效去除多糖多酚对RNA提取的影响，无需苯酚、氯仿等试剂，同时采用硅基质膜吸附RNA进行纯化，提取的总RNA纯度高，无基因组、蛋白质和其它杂质的污染,可用于Real Time RT-PCR、RT-PCR 、Northern Blot、Dot Blot、和体外翻译等多种下游实验。 RNA得率
自备试剂：β-巯基乙醇、无水乙醇（新开封或提取RNA专用） 实验前准备及重要注意事项 1．预防RNase污染，应注意以下几方面： 1）使用无RNase的塑料制品和枪头。 2）操作人员戴一次性口罩和手套，实验过程中要勤换手套。 2．提取的样品避免反复冻融，否则影响RNA提取得率和质量。 3．Buffer RLS如果产生沉淀，请加热使其溶解后室温放置。 4．Buffer RLS在使用前请加入β-巯基乙醇，1 ml Buffer RLS加20 μl β-巯基乙醇。加入 β-巯基乙醇的Buffer RLS室温可保存1个月。 5．第一次使用Buffer RW2前应按照试剂瓶标签的说明加入无水乙醇。 操作步骤 1．匀浆处理：取50-100mg植物组织在液氮中迅速研磨成粉末，加入500 μl Buffer RLS （使用前请先检查是否加入β-巯基乙醇），立即涡旋剧烈震荡混匀。 注意：对于含水量极其丰富的材料，如西瓜果肉，西红柿，梨果肉等，可以适当多加入些材料，最多可增加至200mg；对于富含淀粉的样本或成熟叶片，可适当增加Buffer RLS的用量，最多可增加至700μl。 2．4°C 12,000 rpm（~13,400×g）离心2分钟。 3．将上清液转入已装入收集管的过滤柱（Spin Columns FS）中，4°C 12,000 rpm离心1分钟，小心吸取收集管中的上清并转移至新的RNase-Free 离心管（自备）中， 枪头尽量避免接触收集管中的细胞碎片沉淀。 4．缓慢加入0.5倍上清体积的无水乙醇，混匀（此时可能会出现沉淀），将得到的溶液和沉淀一起转入已装入收集管的吸附柱（Spin Columns RM）中，若一次不能将 全部溶液加入吸附柱中，请分两次转入。4°C 12,000 rpm离心1分钟，弃废液，将吸附柱重新放回收集管中。 5．向吸附柱RM中加入350 μl Buffer RW1，4°C 12,000 rpm离心1分钟，弃废液，将吸附柱重新放回收集管中。 6．配制DNase I 混合液：取52 μl RNase-Free Water，向其中加入8 μl 10×Reaction Buffer和20 μl DNase I（1 U/μl），混匀， 配制成终体积为80 μl的反应液。 7．向吸附柱中直接加入80 µl DNase I 混合液，20-30℃孵育15分钟。 8．向吸附柱RM中加入350 μl Buffer RW1，4°C 12,000 rpm离心1分钟，弃废液，将吸附柱重新放回收集管中。 9．向吸附柱RM中加入500 μl Buffer RW2（使用前检查是否加入无水乙醇）, 4°C 12,000 rpm 离心1 分钟，弃废液，将吸附柱重新放回收集管中。 10．重复步骤9。 11．4°C 12,000 rpm离心2分钟。 注意：这一步的目的是将吸附柱中残余乙醇去除，乙醇残留会影响后续的酶促反应(酶切、PCR等)。 12．将吸附柱RM装入新的RNase-Free Centrifuge Tubes（1.5 ml）中，向吸附膜的中间部位悬空滴加30-50 μl RNase-Free Water，室温放置2分钟，4°C 12,000 rpm离 心1分钟，得到的RNA溶液保存在-70℃，防止降解。 注意：1） RNase-Free Water体积不应小于30 μl，体积过小影响回收率。 2） 如果要提高RNA的产量，可用30-50 μl新的RNase-Free Water重复步骤12。 3） 如果要提高RNA浓度，可将得到的溶液重新加入到吸附柱中，重复步骤12。
Product Introduction This kit is suitable for extracting RNA from a wide range of plants, even from plants rich in polysaccharides and polyphenols, high quality RNA can be successfully extracted, such as rice leaves, wheat leaves, corn leaves, tobacco leaves, pine needles, ginkgo leaves, poplar leaves, pomegranate leaves, holly leaves, apples, peaches, pears, tomatoes, cherries, apricots, bananas, grapes, loquats, cinnamon rinds, cinnamon pulp, lychee fruit rinds, lychee pulp, soybean, peanut, corn, potato tuber, moonflower petal, pomegranate petal, shiitake mushroom, flat mushroom and other samples. The unique lysate formula can rapidly inactivate the RNA enzyme in the cell, effectively remove the effect of polysaccharide and polyphenol on RNA extraction, without the need for phenol, chloroform and other reagents, while using silicon matrix membrane adsorption of RNA for purification, the total RNA extracted is highly pure, without the contamination of genomes, proteins and other impurities, and can be used for Real Time RT-PCR, RT-PCR, It can be used for Real Time RT-PCR, RT-PCR, Northern Blot, Dot Blot, in vitro translation and other downstream experiments. Self-contained reagents: β-mercaptoethanol, anhydrous ethanol (freshly opened or for RNA extraction) Pre-experiment Preparation and Important Notes 1. To prevent RNase contamination, attention should be paid to the following aspects: 1) Use RNase-free plastics and tips. (2) Operators wear disposable masks and gloves, and change gloves diligently during the experiment. 2. Avoid repeated freezing and thawing of the extracted samples, otherwise it will affect the rate and quality of RNA extraction. 3. If Buffer RLS produces a precipitate, heat to dissolve it and leave at room temperature. 4. Please add β-mercaptoethanol to Buffer RLS before use, add 20μl β-mercaptoethanol to 1ml Buffer RLS. Buffer RLS with β-mercaptoethanol can be stored for 1 month at room temperature. 5. Anhydrous ethanol should be added according to the instructions on the reagent bottle label before using Buffer RW2 for the first time. Operation steps 1. Homogenization: Take 50-100mg of plant tissue and quickly grind it into powder in liquid nitrogen, add 500μl of Buffer RLS (please check whether β-mercaptoethanol is added before use), and immediately mix it by vortexing with vigorous shaking. Note: For materials that are extremely rich in water content, such as watermelon pulp, tomato, pear pulp, etc., more material can be added appropriately, up to 200 mg; for starch-rich samples or mature leaves, the amount of Buffer RLS can be increased appropriately, up to 700 μl. 2. Centrifuge at 12,000 rpm (~13,400 x g) for 2 min at 4°C. 3. Transfer the supernatant into the filter columns (Spin Columns FS) that have been loaded into the collection tubes, centrifuge at 12,000 rpm at 4°C for 1 minute, carefully aspirate the supernatant in the collection tubes and transfer it to new RNase-Free centrifugation tubes (self-provided), avoiding the tip of the gun from touching the cell debris precipitation in the collection tubes as much as possible. 4. Slowly add 0.5 times the volume of the supernatant in anhydrous ethanol, mix well (a precipitate may appear), and transfer the resulting solution together with the precipitate to a Spin Columns RM in a collection tube, or in two batches if you cannot add all of the solution at once. centrifuge the column for 1 minute at 12,000 rpm at 4°C. Dispose of the spent solution and place the column back into the collection tube. Centrifuge at 12,000 rpm for 1 minute at 4°C, discard the spent solution and return the column to the collection tube. 5. Add 350 μl of Buffer RW1 to the adsorbent column RM, centrifuge at 12,000 rpm at 4°C for 1 min, discard the waste solution and put the adsorbent column back into the collection tube. 6. Preparation of DNase I mixture: Take 52μl of RNase-Free Water, add 8μl of 10×Reaction Buffer and 20μl of DNase I (1U/μl) to it, mix well, and prepare a final volume of 80μl of reaction solution. 7. Add 80µl of DNase I mixture directly to the adsorption column and incubate at 20-30°C for 15 minutes. 8. Add 350 μl of Buffer RW1 to the adsorbent column RM, centrifuge at 12,000 rpm at 4°C for 1 min, discard the waste solution and put the adsorbent column back into the collection tube. 9. Add 500 μl of Buffer RW2 to the adsorbent column RM (check that anhydrous ethanol is added before use), centrifuge at 12,000 rpm for 1 minute at 4°C, discard the waste solution and put the adsorbent column back into the collection tube. 10. Repeat step 9. 11. Centrifuge at 12,000 rpm for 2 minutes at 4°C. Note: The purpose of this step is to remove residual ethanol from the adsorption column; ethanol residue can interfere with subsequent enzymatic reactions (zymography, PCR, etc.). 12. Load the adsorption column RM into new RNase-Free Centrifuge Tubes (1.5 ml), add 30-50 μl of RNase-Free Water dropwise to the middle part of the adsorption membrane overhang, leave it at room temperature for 2 min, and centrifuge at 12,000 rpm at 4°C for 1 min, and store the resulting RNA solution at -70°C to prevent degradation. Note: 1) The volume of RNase-Free Water should not be less than 30 μl, too small volume affects the recovery rate. 2) If you want to increase the RNA yield, repeat step 12 with 30-50 μl of fresh RNase-Free Water. 3) If the RNA concentration is to be increased, the resulting solution can be reintroduced into the adsorption column and step 12 repeated. |
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