產(chǎn)品名稱

人PPP1R2 (Protein phosphatase 1抑制劑subunit 2) knockout HeLa cell line

Parental Cell Line

HeLa

Organism

Human

Mutation description

Knockout achieved by using CRISPR/Cas9, Homozygous: Insertion of the selection cassette in exon 1

Passage number

<20

Knockout validation

Sanger Sequencing

Biosafety level

2

常規(guī)說明

Recommended control: Human wild-type HeLa cell line (ab255928). Please note a wild-type cell line is not automatically included with a knockout cell line order, if required please add recommended wild-type cell line at no additional cost using the code WILDTYPE-TMTK1.

Cryopreservation cell medium: Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.

Culture medium: DMEM (High Glucose) + 10% FBS

Initial handling guidelines: Upon arrival, the vial should be stored in liquid nitrogen vapor phase and not at -80°C. Storage at -80°C may result in loss of viability.

1. Thaw the vial in 37°C water bath for approximately 1-2 minutes.

2. Transfer the cell suspension (0.8 mL) to a 15 mL/50 mL conical sterile polypropylene centrifuge tube containing 8.4 mL pre-warmed culture medium, wash vial with an additional 0.8 mL culture medium (total volume 10 mL) to collect remaining cells, and centrifuge at 201 x g (rcf) for 5 minutes at room temperature. 10 mL represents minimum recommended dilution. 20 mL represents maximum recommended dilution.

3. Resuspend the cell pellet in 5 mL pre-warmed culture medium and count using a haemocytometer or alternative cell counting method. Based on cell count, seed cells in an appropriate cell culture flask at a density of 2x104 cells/cm2. Seeding density is given as a guide only and should be scaled to align with individual lab schedules.

4. Incubate the culture at 37°C incubator with 5% CO2. Cultures should be monitored daily.

Subculture guidelines:

All seeding densities should be based on cell counts gained by established methods.

A guide seeding density of 2x104 cells/cm2 is recommended.

A partial media change 24 hours prior to subculture may be helpful to encourage growth, if required.

Cells should be passaged when they have achieved 80-90% confluence.

This product is subject to limited use licenses from The Broad Institute, ERS Genomics Limited and Sigma-Aldrich Co. LLC, and is developed with patented technology. For full details of the licenses and patents please refer to our limited use license and patent pages.

艾普蒂生物科技（ipodix）多年來致力于重組蛋白表達與純化的相關(guān)研究，每年可完成上百組蛋白的表達與純化服務(wù)，擁有完整的蛋白純化體系，可根據(jù)客戶的不同需求提供多元化的蛋白純化方案。我們可為您提供定制的蛋白純化服務(wù)包括，有/無標簽重組蛋白，天然蛋白的純化分離等。根據(jù)蛋白性質(zhì)設(shè)計親和純化、離子交換、分子篩、疏水等多種蛋白純化實驗方案，獲得的高純度蛋白質(zhì)，可根據(jù)客戶需求一對一進行個性化方案定制，能夠滿足蛋白質(zhì)折疊、重結(jié)晶等一系列精細實驗要求。

重組蛋白純化：

重組蛋白分離純化一般分為三種：含標簽的重組蛋白純化、不含標簽的重組蛋白純化以及包涵體變性與復(fù)性純化。

含標簽的重組蛋白純化：組氨酸標簽（His-tag）是使用范圍較為廣泛的一種蛋白純化標簽，其純化的原理是組氨酸殘基上的咪唑基團可與層析介質(zhì)中的金屬離子螯合，再通過提高緩沖液中咪唑濃度進行競爭性洗脫達到純化效果，這種純化標簽的有點在于His-Tag的分子質(zhì)量非常小且?guī)缀醪粫δ康牡鞍妆旧淼纳飳W(xué)功能及特性產(chǎn)生影響，因此帶有His-Tag的蛋白純化后可直接作為抗原免疫動物制備抗體并不會影響抗體的特異性。艾普蒂生物還提供其他多種類型標簽的蛋白純化服務(wù)，如GST-Tag、SUMO-Tag、MBP-Tag等。

不含標簽的重組蛋白純化：不含純化標簽的重組蛋白我們會根據(jù)客戶的樣品及需求定制方案，一般會采用抗體親和純化方法，利用靶向配體能與目的蛋白特異性結(jié)合的特性將目的蛋白分離出來；或選用分子篩進行分離純化。

包涵體變性與復(fù)性純化：將以包涵體形式存在的蛋白進行變復(fù)性操作，根據(jù)蛋白質(zhì)不同的性質(zhì)，可以選擇尿素、鹽酸胍、去垢劑、極端pH等，以保證獲得單體狀態(tài)的變性蛋白。

艾普蒂生物提供的蛋白分離純化流程：

服務(wù)優(yōu)勢：

--純化方式多元化，適合不同性質(zhì)的重組蛋白；

--專業(yè)技術(shù)團隊為您量身定制純化方案，獲得能夠滿?下游實驗的?純度蛋?；

--提供全面的蛋白純化分析報告；