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AB255410

Human MSH6 knockout HeLa cell line

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MSH6 KO cell line available to order. KO validated by Western blot. Free of charge wild type control available. Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 4. To order both knockout and wild-type control cells: select '2 x 1000000 Cells/vial'. To order only knockout cells: select '1000000 Cells/vial'.
3 Images
Western blot - Human MSH6 knockout HeLa cell line (AB255410)
  • WB

Lab

Western blot - Human MSH6 knockout HeLa cell line (AB255410)

Lanes 1- 2 : Merged signal (red and green). Green - ab14204 observed at 160 kDa. Red - Anti-alpha Tubulin antibody [EP1332Y] - Microtubule Marker (ab52866) observed at 50 kDa.

ab14204 was shown to react with MSH6 in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab255410 (knockout cell lysate ab263763) was used. Wild-type HeLa and MSH6 knockout HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab14204 and Anti-alpha Tubulin antibody [EP1332Y] - Microtubule Marker (ab52866) overnight at 4° at a 1 in 500 dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye®800CW) preadsorbed (ab216772) and Goat Anti-Rabbit IgG H&L (IRDye®680RD) preadsorbed (ab216777) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-MSH6 antibody [44] (<a href='/en-us/products/primary-antibodies/msh6-antibody-44-ab14204'>ab14204</a>) at 1/500 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

MSH6 knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human MSH6 knockout HeLa cell line (ab255410)

Predicted band size: 153 kDa

Observed band size: 160 kDa

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Western blot - Human MSH6 knockout HeLa cell line (AB255410)
  • WB

Lab

Western blot - Human MSH6 knockout HeLa cell line (AB255410)

Lanes 1- 2 : Merged signal (red and green). Green - ab92471 observed at 160 kDa. Red - Anti-Vinculin antibody [VIN-54] observed at 124 kDa.

ab92471 was shown to react with MSH6 in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab255410 (knockout cell lysate ab263763) was used. Wild-type HeLa and MSH6 knockout HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab92471 and Anti-Vinculin antibody [VIN-54] overnight at 4° at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye®800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye®680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-MSH6 antibody [EPR3945] (<a href='/en-us/products/primary-antibodies/msh6-antibody-epr3945-ab92471'>ab92471</a>) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

MSH6 knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human MSH6 knockout HeLa cell line (ab255410)

Predicted band size: 153 kDa

Observed band size: 160 kDa

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Sanger Sequencing - Human MSH6 knockout HeLa cell line (AB255410)
  • Sanger seq

Unknown

Sanger Sequencing - Human MSH6 knockout HeLa cell line (AB255410)

Homozygous : 1 bp insertion in exon 4.

Key facts

Cell type

HeLa

Species or organism

Human

Tissue

Cervix

Form

Liquid

form

Knockout validation

Sanger Sequencing,Western blot

Mutation description

Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 4

Disease

Adenocarcinoma

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Primary Antibodies

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-MSH6 antibody [EPR3945] (AB92471)

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Primary Antibodies

AB179483

Anti-HIF-1 alpha antibody [EPR16897]

RabMAb|Advanced Validation|Recombinant|KO Validated

Immunocytochemistry/ Immunofluorescence - Anti-HIF-1 alpha antibody [EPR16897] (AB179483)

  • 4
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Reactivity data

{ "title": "Reactivity Data", "filters": { "stats": ["", "Reactivity", "Dilution Info", "Notes"] }, "values": { "WB": { "reactivity":"TESTED_AND_REACTS", "dilution-info":"", "notes":"<p></p>" } } }
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Product details

We will provide viable cells that proliferate on revival.

Western blot data indicates that the CRISPR gene edit may have resulted in a truncation of the protein of interest. Please see data images.

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.

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What's included?

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Properties and storage information

Gene name
MSH6
Gene editing type
Knockout
Gene editing method
CRISPR technology
Knockout validation
Sanger Sequencing, Western blot
Zygosity
Homozygous
Shipped at conditions
Dry Ice
Appropriate short-term storage conditions
-196°C
Appropriate long-term storage conditions
-196°C
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Handling procedures

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 seed all remaining cells into a T25.
4. Incubate the culture at 37°C incubator with 5% CO2. Check the culture one day after revival and continue to check until 80% confluent. Media change can be given if needed.
5. Once confluent passage into an appropriate 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. 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.
  • Cells should be passaged when they have achieved 80-90% confluence.
Culture medium

DMEM (High Glucose) + 10% FBS

Cryopreservation medium

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

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Supplementary information

This supplementary information is collated from multiple sources and compiled automatically.

MSH6 also known as MutS Homolog 6 is a DNA repair protein that plays a role in the mismatch repair (MMR) system. It has a molecular mass of approximately 136 kDa. MSH6 forms a heterodimer with MSH2 called MutSα and this complex identifies base-pair mismatches and insertion-deletion loops during DNA replication. It is expressed in various tissues throughout the body and high levels are often found in proliferative tissues where active DNA replication occurs.
Biological function summary

MSH6 functions as part of the MMR complex which is essential for maintaining genomic stability. The MutSα complex where MSH6 pairs with MSH2 operates along with other proteins in the MMR pathway to correct DNA replication errors. MSH6 is also known to interact with PCNA a DNA polymerase processivity factor which facilitates its role in the repair process.

Pathways

MSH6 participates prominently in the DNA mismatch repair pathway. This pathway is critical for correcting DNA errors and preventing mutations during replication. In association with MLH1-PMS2 (MutLα complex) MSH6 ensures that DNA integrity is preserved. Additionally MSH6 is involved in the base excision repair (BER) pathway where it collaborates with other repair proteins to fix small base lesions.

MSH6 has a significant connection with Lynch syndrome also known as hereditary nonpolyposis colorectal cancer (HNPCC). This condition is characterized by germline mutations in MMR genes including MSH6 leading to increased cancer risk particularly in the colon. Moreover alterations in MSH6 can contribute to microsatellite instability a feature seen in certain types of endometrial cancer. Mutations in MSH2 often accompany MSH6 mutations in these disorders further impacting the MMR pathway's efficiency.
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Quality control

STR analysis

CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX

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Cell culture

Biosafety level

EU: 2 US: 2

Adherent/suspension

Adherent

Gender

Female

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Product protocols

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Target data

See full target information MSH6
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Product promise

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For full details, please see our Terms & Conditions

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