基本描述

英文名称

PurePlasmid Mini Kit

储存温度

室温

运输条件

常规运输

产品介绍

产品内容

P666142	Component	200 T	Storage
P666142A	Buffer P1	60 mL	RT
P666142B	Buffer P2	60 mL	RT
P666142C	Buffer N3	80 mL	RT
P666142D	Buffer PB	35 mL	RT
P666142E	Buffer PW (concentrate)	25 mL	RT
P666142F	Buffer EB	30 mL	RT
P666142G	RNase A (10 mg/mL)	600 μL	RT
P666142H	Spin Columns DM with Collection Tubes	200 EA	RT

产品简介

本试剂盒适合提取1-5 ml菌液，在碱裂解法裂解细胞的基础上，采用独特的硅基质膜吸附技术和试剂配方，通过离心吸附柱在高盐状态下高效专一的结合溶液中的质粒DNA，每个吸附柱最高可吸附30 μg的质粒DNA，并最大限度的去除蛋白质、基因组、RNA和其他杂质。得到的质粒DNA可直接用于细胞转染、PCR、酶切、测序、连接等生物学实验。

自备试剂：无水乙醇。

实验前准备及重要注意事项

1. 所有组分可在干燥、室温（15-30℃）环境稳定保存1年，将吸附柱置于2-8℃可保存更长时间，加入RNase A的Buffer P1 置于2-8℃可稳定保存6个月。

2. 第一次使用前，将RNase A溶液全部加入到Buffer P1中，混匀，置于2–8℃保存，使用前需在室温中放置一段时间，恢复至室温后使用。

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

4. 使用前若发现Buffer P2、Buffer N3、Buffer PB有沉淀，可在37℃水浴几分钟，即可恢复澄清（请勿剧烈晃动Buffer P2）。

5. 注意不要直接接触Buffer P2 、Buffer N3和 Buffer PB，使用后应立即盖紧盖子。

6．提取质粒的量和纯度与细菌培养浓度、菌株种类、质粒大小、质粒拷贝数等因素有关。

操作步骤

1. 取1-5 ml过夜培养的菌液加入离心管（自备）中，13,000 rpm（~16,200×g）离心30 秒收集菌体沉淀，尽量吸弃上清。

2. 向留有菌体沉淀的离心管中加入250 μl Buffer P1（请先检查是否已加入RNase A），使用移液器或涡旋振荡器充分混匀，悬浮菌体沉淀。

注意：如果菌块未彻底混匀，将会影响裂解效果，导致提取量和纯度偏低。

3. 向离心管中加入250 μl Buffer P2，温和地上下颠倒混匀4-6次，充分混匀使菌体裂解，此时溶液应变得清亮粘稠。

注意：温和混匀，不要剧烈震荡，以免打断基因组DNA，造成提取的质粒中混有基因组DNA片段。此步骤所用时间应不超过5分钟，避免质粒受到破坏。

4. 向离心管中加入350 μl Buffer N3，立即温和地上下颠倒混匀8-10次，充分混匀，此时应出现白色絮状沉淀。13,000 rpm离心5分钟。

注意：Buffer N3加入后应立即混匀，避免产生局部沉淀。

5. 将步骤4中所得的上清液转移到已装入收集管的吸附柱（Spin Columns DM）中，13,000 rpm离心30秒钟，倒掉收集管中的废液，将吸附柱重新放回收集管中。

6. 向吸附柱中加入150 μl Buffer PB，13,000 rpm离心30秒。

7. 向吸附柱中加入400 μl Buffer PW（请先检查是否已加入无水乙醇），13,000 rpm离心1分钟，倒掉收集管中的废液。

8. 将吸附柱置于一个新的离心管（自备）中，向吸附膜的中间部位加入50-100 μl Buffer EB，室温放置2分钟，13,000 rpm离心1分钟，将质粒溶液收集到离心管中。-20℃ 保存质粒。

注意：1）为了增加质粒的回收效率，可将得到的溶液重新加入到吸附柱中，室温放置2分钟，13,000 rpm离心1分钟，将质粒溶液收集到离心管中。

2) 质粒拷贝数较低或>10 kb时， Buffer EB在65-70℃水浴预热，可以增加提取效率。

Product content

P666142	Component	200 T	Storage
P666142A	Buffer P1	60 mL	RT
P666142B	Buffer P2	60 mL	RT
P666142C	Buffer N3	80 mL	RT
P666142D	Buffer PB	35 mL	RT
P666142E	Buffer PW (concentrate)	25 mL	RT
P666142F	Buffer EB	30 mL	RT
P666142G	RNase A (10 mg/mL)	600 μL	RT
P666142H	Spin Columns DM with Collection Tubes	200 EA	RT

Product Introduction

This kit is suitable for extracting 1-5 ml of bacterial solution. Based on the lysis of cells by alkaline lysis method, it adopts a unique silica matrix membrane adsorption technology and reagent formulation, and efficiently and exclusively binds plasmid DNA in solution by centrifugal adsorption columns in a high-salt state, and each adsorption column can adsorb a maximum of 30 μg of plasmid DNA, and removes proteins, genomes, RNAs, and other impurities to the greatest extent possible. The plasmid DNA obtained can be directly used for cell transfection, PCR, digestion, sequencing, ligation and other biological experiments.

Self-contained reagent: anhydrous ethanol.

Pre-experiment Preparation and Important Notes

1. All components can be stably stored in dry, room temperature (15-30℃) environment for 1 year, the adsorption column can be stored at 2-8℃ for a longer period of time, and Buffer P1 with RNase A can be stably stored at 2-8℃ for 6 months.

2. Before the first use, add all the RNase A solution into Buffer P1, mix well, and store it at 2-8°C. Before use, leave it at room temperature for a period of time, and then use it after recovering to room temperature.

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

4. If precipitation is found in Buffer P2, Buffer N3, or Buffer PB before use, the clarification can be restored by water bath at 37℃ for a few minutes (please do not shake Buffer P2 violently).

5. Be careful not to touch Buffer P2, Buffer N3 and Buffer PB directly, and tighten the lid immediately after use.

6. The amount and purity of extracted plasmid is related to the concentration of bacterial culture, strain type, plasmid size, plasmid copy number and other factors.

Procedure

1. Take 1-5 ml of the overnight culture and add it to a centrifuge tube (self-prepared), centrifuge for 30 seconds at 13,000 rpm (~16,200×g) to collect the bacterial precipitate, and discard the supernatant as much as possible.

2. Add 250 μl of Buffer P1 to the centrifuge tube with the bacterial precipitate (please check that RNase A has been added first), mix well using a pipette or vortex shaker, and suspend the bacterial precipitate.

Note: If the bacterial mass is not thoroughly mixed, it will affect the lysis effect, resulting in low extraction and purity.

3. Add 250μl of Buffer P2 to the centrifuge tube and mix gently up and down 4-6 times, mixing well to lyse the organisms, at which point the solution should become clear and viscous.

Note: Mix gently, do not shake vigorously to avoid interrupting the genomic DNA and causing the extracted plasmid to be mixed with genomic DNA fragments. This step should take no more than 5 minutes to avoid damage to the plasmid.

4. Add 350 μl of Buffer N3 to the centrifuge tube and immediately mix gently up and down for 8-10 times, mixing well so that a white flocculent precipitate should appear. centrifuge at 13,000 rpm for 5 minutes.

Note: Buffer N3 should be mixed immediately after addition to avoid localized precipitation.

5. Transfer the supernatant obtained in step 4 to the Spin Columns DM that have been loaded into the collection tube, centrifuge at 13,000 rpm for 30 seconds, pour off the waste liquid from the collection tube, and place the column back into the collection tube.

6. Add 150 μl Buffer PB to the adsorption column and centrifuge at 13,000 rpm for 30 seconds.

7. Add 400 μl Buffer PW to the adsorption column (please check that anhydrous ethanol has been added first), centrifuge at 13,000 rpm for 1 minute, and pour off the waste liquid in the collection tube.

8. Place the adsorbent column in a new centrifuge tube (supplied), add 50-100 μl Buffer EB to the middle of the adsorbent membrane, leave it at room temperature for 2 minutes, centrifuge at 13,000 rpm for 1 minute, and collect the plasmid solution into the centrifuge tube. -The plasmid solution was collected into the centrifuge tube.

Note: 1) To increase the recovery efficiency of the plasmid, the resulting solution can be reintroduced into the adsorbent column, left at room temperature for 2 minutes, centrifuged at 13,000 rpm for 1 minute, and the plasmid solution collected into a centrifuge tube.

2) For low plasmid copy number or >10 kb, Buffer EB is preheated at 65-70°C in a water bath to increase extraction efficiency.

安全和危险性（GHS）

SDS英文版

质量标准

质量标准

p666142_pureplasmid mini kit.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

计算溶液所需的质量、体积或浓度。

质量

=

浓度

x

体积

x

分子量

g/mol

稀释计算器

计算配制储备溶液所需的稀释度。

浓度1（C1）

x

体积 1 (V1)

=

浓度 2 (C2)

x

体积 2 (V2)

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。

体积（添加到小瓶中）

=

质量（小瓶）

÷

所需的复溶浓度

高纯度质粒小提试剂盒

库存信息

基本描述

安全和危险性（GHS）

质量标准

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