计算溶液所需的质量、体积或浓度。
粪便总RNA提取试剂盒 (DNase I)
有货
为什么选择此级别
BioReagent,分子生物学级 ，适用于对基线干扰要求严格的色谱和分析工作流程。
储存与运输
2-8°C储存,室温。低温运输 。请查阅批次 COA 获取详细规格。
质量文档
SDS、COA、产品数据表及规格说明书均可下载。可通过批号查询获取批次 COA。
文献证明
在色谱分析、有机合成和交叉偶联反应领域已被 0 篇同行评审文献引用。
概述
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，可从粪便样本中快速提取总RNA。提取的总RNA纯度极高，没有蛋白质和其他污染，可以直接用于Northern杂交、斑点杂交、mRNA纯化、体外翻译、RNA分解酶的保护分析、RT-PCR、RT-qPCR和构建cDNA文库等各种分子生物学实验。
注意事项
1. 操作人员应戴一次性口罩和手套，实验过程中要勤换手套，并使用无RNase的塑料制品及枪头，避免RNase污染。
2. DNase I 储存液的配制：用1 mL注射器将550 μL RNase-Free ddH₂O注入装有DNase I 干粉的瓶中，轻柔混匀至溶解，分装后-30~-15℃储存（可保存9个月），每次根据用量需求取出。配制的DNase I 工作液置于2-8℃储存（可保存42天），避免反复冻融。
操作步骤
使用前准备：在12 mL Buffer RW2 中加入48 mL的无水乙醇。
1. 在匀浆管 A 中加入 660 μL Buffer ST1，称取 180–220 mg 粪便样本至匀浆管 A 中，盖上管盖，上下剧烈摇晃约 20 次。加入 200 μL Buffer ST2，将匀浆管放入 25℃、1500 rpm 的恒温混匀仪上振荡混匀 10–15 min，使其充分裂解，12,000 rpm（13,680 × g）离心 5 min。
注意：
若使用组织研磨仪：研磨 30–60 sec，共 2 次（调整机器 Hz，防止离心管破碎），然后 12,000 rpm（13,400 × g）离心 5 min。
样本称量调整：含杂质较多的粪便样本：避免称入未消化的食物残渣（如谷物、果皮种子等）。较干燥的粪便样本（如兔、小鼠）：称取 30–80 mg。油脂、多糖或蛋白多的粪便样本：称取 50–100 mg。
新鲜采集的粪便：可直接提取，或于 2–8℃ 保存不超过 24 h。
- 不能及时提取的样本：冻存于 -20℃ 或 -80℃。提取前在冰上过夜解冻或室温快速解冻，避免反复冻融。
2. 吸取 510 μL 上清至新的离心管中，加入 140 μL Buffer ST3，盖盖，涡旋振荡混匀 5 sec。12,000 rpm（13,680 × g）离心 3 min。
3. 转移全部上清液至 抑制物清除柱（IR）（清除柱放在收集管中），12,000 rpm（13,680 × g）离心 1 min，弃去清除柱，收集滤液。
4. 向滤液中加入 180 μL Buffer ST3，混匀。将混合液转移至 吸附柱 RA（吸附柱放在收集管中）。12,000 rpm（13,680 × g）离心 1 min。弃废液，将吸附柱放回收集管中。
5. 向吸附柱中加入 500 μL Buffer STW1，12,000 rpm（13,680 × g）离心 30 sec，弃废液，将吸附柱放回收集管中。
6. DNase I 工作液配制：取 10 μL DNase I 储存液 放入新的 1.5 mL RNase-Free 离心管中，加入 70 μL DNase I Buffer，轻柔混匀。
7. 向吸附柱中央加入 80 μL DNase I 工作液，室温放置 15 min。
8. 向吸附柱中加入 500 μL Buffer STW2，12,000 rpm（13,680 × g）离心 30 sec，弃废液，将吸附柱放回收集管中。
9. 向吸附柱中加入 500 μL Buffer STW3，12,000 rpm（13,680 × g）离心 30 sec，弃废液，将吸附柱放回收集管中。
10. 向吸附柱中加入 500 μL Buffer RW2（使用前请加入无水乙醇），12,000 rpm（13,680 × g）离心 30 sec，弃废液，将吸附柱放回收集管中。
11. 12,000 rpm（13,680 × g）离心 2 min。
12. 将吸附柱转移至一个新的 1.5 mL RNase-Free 离心管中，向吸附膜中央悬空滴加 50–100 μL RNase-Free ddH₂O，室温放置 1 min，12,000 rpm（13,680 × g）离心 30 sec。所得 RNA 可直接用于下游实验，或于 -80℃ 以下保存。
注意：
RNase-Free ddH₂O 体积不应小于 50 μL，体积过小影响回收率。
若要提高 RNA 产量：可将第一次洗脱液重新加入吸附柱再次洗脱，或将 RNase-Free ddH₂O 预热至 65℃ 后再使用。
This kit uses a high-efficiency, nucleic acid‑binding spin column and a unique buffer system to rapidly extract total RNA from fecal samples. The extracted total RNA is of extremely high purity, free of proteins and other contaminants, and can be directly used in various molecular biology experiments such as Northern blot, dot blot, mRNA purification, in vitro translation, RNase protection assay, RT‑PCR, RT‑qPCR, and cDNA library construction.
Precautions
Operators should wear disposable masks and gloves, change gloves frequently during the experiment, and use RNase‑free plasticware and pipette tips to avoid RNase contamination.
Preparation of DNase I stock solution: Use the 1 mL syringe to inject 550 μL of RNase‑Free ddH₂O into the vial containing DNase I dry powder. Mix gently until dissolved. Aliquot and store at –30 to –15°C (stable for 9 months). Thaw only the amount needed for each use. The prepared DNase I working solution can be stored at 2–8°C for up to 42 days; avoid repeated freeze‑thaw cycles.
Operating Procedure
Preparation before use: Add 48 mL of absolute ethanol to 12 mL of Buffer RW2.
1. Add 660 μL Buffer ST1 to a Homogenization Tube A. Weigh 180–220 mg of fecal sample into the tube, close the cap, and shake vigorously up and down about 20 times. Add 200 μL Buffer ST2. Place the tube on a thermomixer at 25°C and 1500 rpm, shake for 10–15 min to ensure thorough lysis. Centrifuge at 12,000 rpm (13,680 × g) for 5 min.
Note:
- If using a tissue homogenizer: Homogenize for 30–60 sec, twice (adjust machine Hz to prevent tube breakage), then centrifuge at 12,000 rpm (13,400 × g) for 5 min.
- Sample weight adjustments:
For fecal samples with many impurities: avoid including undigested food residues (e.g., grains, fruit peels/seeds).
For dry fecal samples (e.g., rabbit, mouse): weigh 30–80 mg.
For fecal samples rich in lipids, polysaccharides, or proteins: weigh 50–100 mg. - Freshly collected feces can be processed immediately or stored at 2–8°C for no more than 24 h.
- If extraction cannot be performed promptly, freeze the sample at –20°C or –80°C. Thaw on ice overnight or rapidly at room temperature before extraction, and avoid repeated freeze‑thaw cycles.
2. Transfer 510 μL of supernatant to a new centrifuge tube. Add 140 μL Buffer ST3, close the cap, and vortex for 5 sec. Centrifuge at 12,000 rpm (13,680 × g) for 3 min.
3. Transfer the entire supernatant to an Inhibitor Removal Column (IR) (placed in a collection tube). Centrifuge at 12,000 rpm (13,680 × g) for 1 min. Discard the column and keep the flow‑through.
4. Add 180 μL Buffer ST3 to the flow‑through and mix. Transfer the mixture to an Adsorption Column RA (placed in a collection tube). Centrifuge at 12,000 rpm (13,680 × g) for 1 min. Discard the flow‑through and place the column back into the collection tube.
5. Add 500 μL Buffer STW1 to the column. Centrifuge at 12,000 rpm (13,680 × g) for 30 sec. Discard the flow‑through and place the column back into the collection tube.
6. Preparation of DNase I working solution: Transfer 10 μL of DNase I stock solution into a new 1.5 mL RNase‑Free centrifuge tube, add 70 μL DNase I Buffer, and mix gently.
7. Add 80 μL of the DNase I working solution to the center of the column membrane. Incubate at room temperature for 15 min.
8. Add 500 μL Buffer STW2 to the column. Centrifuge at 12,000 rpm (13,680 × g) for 30 sec. Discard the flow‑through and place the column back into the collection tube.
9. Add 500 μL Buffer STW3 to the column. Centrifuge at 12,000 rpm (13,680 × g) for 30 sec. Discard the flow‑through and place the column back into the collection tube.
10. Add 500 μL Buffer RW2 (make sure ethanol has been added before use). Centrifuge at 12,000 rpm (13,680 × g) for 30 sec. Discard the flow‑through and place the column back into the collection tube.
11. Centrifuge at 12,000 rpm (13,680 × g) for 2 min.
12. Transfer the column to a new 1.5 mL RNase‑Free centrifuge tube. Add 50–100 μL RNase‑Free ddH₂O to the center of the membrane, drop by drop (do not touch the membrane). Incubate at room temperature for 1 min. Centrifuge at 12,000 rpm (13,680 × g) for 30 sec. The obtained RNA can be used directly in downstream experiments or stored at –80°C or below.
Note:
The volume of RNase‑Free ddH₂O should not be less than 50 μL; smaller volumes reduce recovery.
To increase RNA yield: re‑apply the first eluate to the column for a second elution, or pre‑heat the RNase‑Free ddH₂O to 65°C before use.
规格
别名
柱式粪便RNA提取试剂盒
英文别名
Fecal Total RNA Extraction Kit (DNase I)
规格或纯度
BioReagent,分子生物学级
英文名称
Fecal Total RNA Extraction Kit (DNase I)
应用
RNA Extraction
样品类型
粪便
储存与运输
储存条件
2-8°C储存,室温
运输条件
低温运输
稳定性与储存
各组分在相应的储存条件下保质期15个月。
组分与储存
| F1521385 | Component | 50T | Storage |
| F1521385A | Buffer ST1 | 35 mL | RT. |
| F1521385B | Buffer ST2 | 12 mL | RT. |
| F1521385C | Buffer ST3 | 18 mL | RT. |
| F1521385D | Buffer STW1 | 28 mL | RT. |
| F1521385E | Buffer STW2 | 28 mL | RT. |
| F1521385F | Buffer STW3 | 28 mL | RT. |
| F1521385G | Buffer RW2 | 12 mL | RT. |
| F1521385H | RNase-Free ddH₂O | 15 mL | RT. |
| F1521385I | 匀浆管A | 1 EA×50 | RT. |
| F1521385J | 抑制物清除柱IR (含2mL收集管) | 1 EA×50 | RT. |
| F1521385K | 吸附柱RA (含2mL收集管) | 1 EA×50 | RT. |
| F1521385L | 1mL注射器 | 1 EA | RT. |
| F1521385M | DNase I | 1 EA | 2-8℃. |
| F1521385N | DNase I Buffer | 4 mL | 2-8℃. |
| F1521385O | RNase-Free ddH₂O | 1 mL | 2-8℃. |
F1521385A, F1521385B, F1521385C, F1521385D, F1521385E, F1521385F, F1521385G, F1521385H, F1521385I, F1521385J, F1521385K, F1521385L：室温保存，15个月。
F1521385M, F1521385N, F1521385O：2-8℃保存，15个月。
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| 批号(Lot Number) | 证书类型 | 货号 |
|---|---|---|
 ZJ26F0636302 |
分析证书 | F1521385 |
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