基本描述

英文名称

RNApure Bacteria Kit（DNase I）

储存温度

室温,-20°C储存,避免反复冻融

运输条件

超低温冰袋运输

产品介绍

产品内容

R669890	Component	50 T	Storage
R669890A	DNase I	1000 U	-20℃. Avoid freeze/thaw cycle.
R669890B	10×Reaction Buffer	1mL	-20℃. Avoid freeze/thaw cycle.
R669890C	Buffer RL	35 mL	RT
R669890D	Buffer RW1	40 mL	RT
R669890E	Buffer RW2 (concentrate)	11 mL	RT
R669890F	RNase-Free Water	10 mL	RT
R669890G	Spin Columns FL with Collection Tubes	50 sets	RT
R669890H	Spin Columns RM with Collection Tubes	50 sets	RT
R669890I	RNase-Free Centrifuge Tubes (1.5 mL)	100 EA	RT

产品简介

本试剂盒采用高效、专一结合核酸的离心吸附柱和独特的缓冲系统，可从细菌或培养的动物细胞中快速提取总RNA。30-40分钟内即可完成反应，提取的总RNA纯度极高，没有蛋白质和其他污染，适用于RT-PCR、Real-Time RT-PCR、芯片分析、体外翻译等实验。

自备试剂：Lysozyme、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，避免交叉污染。

2）配制溶液应使用无RNase的水。

3）操作人员戴一次性口罩和手套，实验过程中要勤换手套。

2. Buffer RL在使用前请加入β-巯基乙醇至终浓度为1%，如1 ml Buffer RL加10 μl β-巯基乙醇。加入β-巯基乙醇的Buffer RL 4℃可保存1个月，如出现沉淀，请加热溶解后使用。

3. 第一次使用前应按照试剂瓶标签的说明先在Buffer RW2中加入无水乙醇。

4. 如未特殊说明，所有离心步骤均在室温下进行，且所有操作步骤动作要迅速。

操作步骤

1．4℃ 12,000 rpm（~13,400×g）离心2分钟收集菌体（菌体量最大不超过1×109），小心去除所有上清。

注意：上清如果有残留，会影响后续的消化过程。

2. 用含有Lysozyme的100 μl TE缓冲液彻底重悬菌体，室温孵育。具体配方和孵育时间如下：

/	TE缓冲液中Lysozyme的终浓度	孵育时间
G-细菌	400μg/ml	3-5分钟
G+细菌	3mg/ml	5-10分钟

3. 加入350μl Buffer RL（使用前请检查是否加入β-巯基乙醇），涡旋震荡混匀（此步骤可能出现不溶性沉淀），将溶液与沉淀全部加入到已装入收集管的过滤柱（Spin Columns FL）中，12,000 rpm离心2分钟。

4. 向上步得到的滤液中加入250μl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，12,000 rpm 离心1分钟，弃掉废液，将吸附柱重新放回收集管中。

5. 向吸附柱中加入350 μl Buffer RW1，12,000 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. 向吸附柱中加入350 μl Buffer RW1，12,000 rpm离心1分钟，弃废液，将吸附柱重新放回收集管中。

9. 向吸附柱中加入500 μl Buffer RW2（使用前检查是否加入无水乙醇）, 12,000 rpm离心1分钟,弃废液。

10. 重复步骤9。

11. 将吸附柱放回收集管中，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.将吸附柱装入新的RNase-Free的收集管中，向吸附膜的中间加入30-50 μl RNase-Free Water，室温放置1分钟，12,000 rpm离心1分钟，收集RNA溶液，-70℃保存RNA，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。

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。

Products

R669890	Component	50 T	Storage
R669890A	DNase I	1000 U	-20℃. Avoid freeze/thaw cycle.
R669890B	10×Reaction Buffer	1mL	-20℃. Avoid freeze/thaw cycle.
R669890C	Buffer RL	35 mL	RT
R669890D	Buffer RW1	40 mL	RT
R669890E	Buffer RW2 (concentrate)	11 mL	RT
R669890F	RNase-Free Water	10 mL	RT
R669890G	Spin Columns FL with Collection Tubes	50 sets	RT
R669890H	Spin Columns RM with Collection Tubes	50 sets	RT
R669890I	RNase-Free Centrifuge Tubes (1.5 mL)	100 EA	RT

Products

This kit adopts centrifugal adsorption columns with high efficiency and specificbinding of nucleic acids and unique buffer system, which can rapidly extract totalRNA from bacteria or cultured animal cells.The reaction can be completed in 30-40minutes, and the extracted total RNA is extremely pure and free of protein and othercontaminants, which is suitable for RT-PCR, Real-Time RT-PCR, microarray analysis,in vitro translation and other experiments. Self-contained reagents: Lysozyme, β-mercaptoethanol, anhydrous ethanol (freshlyopened 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 to avoid cross-contamination.

2) RNase-free water should be used to prepare the solution.

3) Operators wear disposable masks and gloves, and change gloves diligently duringthe experiment.

2. Add β-mercaptoethanol to Buffer RL before use to reach a final concentrationof 1%, e.g., add 10 μl of β-mercaptoethanol to 1 ml of Buffer RL. Buffer RL withβ-mercaptoethanol can be stored at 4℃ for 1 month, if precipitation occurs, pleaseheat to dissolve and use.

3. Anhydrous ethanol should be added to Buffer RW2 before first use according tothe instructions on the reagent bottle label.

4. All centrifugation steps are carried out at room temperature if not otherwisespecified, and all steps should be performed quickly.

Procedure

1. Centrifuge at 12,000 rpm (~13,400 x g) at 4°C for 2 minutes to collect theorganisms (maximum volume of organisms should not exceed 1 x 109) and carefullyremove all supernatants. Note: Supernatants that leave residues can interfere with the subsequent digestionprocess.

2. Thoroughly resuspend the organisms with 100 μl of TE buffer containing Lysozymeand incubate at room temperature. The specific formulation and incubation time areas follows:

/	The final concentration of Lysozyme in TE buffer	incubation time
G-germ	400μg/ml	3-5min
G+germ	3mg/ml	5-10min

3. Add 350 μl of Buffer RL (check that β-mercaptoethanol has been added beforeuse), vortex and shake to mix (insoluble precipitate may appear in this step), addall of the solution and the precipitate to the filter columns (Spin Columns FL) thathave been loaded into the collection tubes, and centrifuge at 12,000 rpm for 2minutes. 4. Add 250 μl of anhydrous ethanol to the filtrate obtained in the previous stepand mix well (a precipitate may appear at this point). Transfer the resulting solution together with the precipitate to a Spin Columns RM packed in a collectiontube, centrifuge at 12,000 rpm for 1 min, discard the waste solution and put thecolumn back into the collection tube.

5. Add 350 μl Buffer RW1 to the adsorbent column, centrifuge at 12,000 rpm for1min, discard the waste liquid and put the adsorbent column back into the collectiontube.

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 make a finalvolume of 80μl of reaction solution.

7. Add 80µl of DNase I mixture directly to the adsorption column and incubate at20-30°C for 15 minutes.

8. Add 350 μl Buffer RW1 to the adsorbent column, centrifuge at 12,000 rpm for1min, discard the waste liquid and put the adsorbent column back into the collectiontube.

9. Add 500 μl of Buffer RW2 to the column (check that anhydrous ethanol is addedbefore use), centrifuge at 12,000 rpm for 1 min, and discard the waste solution.

10. Repeat step 9.

11. Place the adsorbent column back into the collection tube and centrifuge at 12,000rpm for 2 minutes. Note: The purpose of this step is to remove residual ethanol from the adsorptioncolumn; ethanol residue can interfere with subsequent enzymatic reactions (zymography, PCR, etc.).

12. Load the adsorption column into a new RNase-Free collection tube, add 30-50 μl of RNase-Free Water to the middle of the adsorption membrane, leave it at roomtemperature for 1 minute, centrifuge at 12,000 rpm for 1 minute, collect the RNAsolution, and store the RNA at -70°C to prevent degradation. Note: 1) The volume of RNase-Free Water should not be less than 30 μl, too smallvolume affects the recovery rate. 2) If you want to increase the RNA yield, repeat step 12 with 30-50 μl of freshRNase-Free Water. If the RNA concentration is to be increased, the resulting solution can be reintroduced into the adsorption column and step 12 repeated.

AI解读

安全和危险性（GHS）

SDS英文版

技术规格说明书

技术规格说明书

r669890_rnapure bacteria kit（dnase i）.pdf

质检证书（CoA,COO,BSE/TSE 和分析图谱)

C of A & Other Certificates(BSE/TSE, COO):

输入批号以搜索分析图谱:

溶液计算器

摩尔浓度计算器

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稀释计算器

计算配制储备溶液所需的稀释度。

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浓度 2 (C2)

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复溶计算器

复溶计算器允许您快速计算试剂的体积，以复溶小瓶。只需输入试剂的质量和目标浓度，计算器将确定其余部分。

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所需的复溶浓度

细菌RNA提取试剂盒（DNase I）

库存信息

基本描述

AI解读

安全和危险性（GHS）

技术规格说明书

质检证书（CoA,COO,BSE/TSE 和分析图谱)

溶液计算器

摩尔浓度计算器

稀释计算器

复溶计算器