Decreased COL11a1 Levels Effect Gene Expression in Chondrocytes
Presentation Date
7-2015
Abstract
Collagen XI alpha 1 (COL11a1) is a critical part of the extracellular matrix for chondrocytes. The purpose of this research is to determine if decreased levels of the COL11a1 gene lead to enhanced expression of osteoarthritic factors in chondrocytes. The down regulation of the COL11a1 gene is achieved using a combination of Clustered Regularly Interspaced Short Palindromic Repeats / CRISPR Associated Protein 9 (CRISPR/CAS9) nuclease vector system and Ribonucleic Acid interference (RNAi). CRISPR/CAS9 vectors and RNAi are prepared and transfected into chondrocytes to knockdown the expression of the COL11a1 gene. Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR) is used to verify the successful knockdown of COL11a1 and a Western Blots is used for protein verification. Differential gene expression is determined by comparison of the COL11a1 knockdown chondrocyte model and the normal chondrocyte model. Additionally, activation of cell signaling pathways are determined by Luciferase assays. Down regulation of the COL11a1 gene is expected to enhance the expression of osteoarthritic factors. This data can be used to procure a treatment to combat these osteoarthritic occurrences.
Decreased COL11a1 Levels Effect Gene Expression in Chondrocytes
Collagen XI alpha 1 (COL11a1) is a critical part of the extracellular matrix for chondrocytes. The purpose of this research is to determine if decreased levels of the COL11a1 gene lead to enhanced expression of osteoarthritic factors in chondrocytes. The down regulation of the COL11a1 gene is achieved using a combination of Clustered Regularly Interspaced Short Palindromic Repeats / CRISPR Associated Protein 9 (CRISPR/CAS9) nuclease vector system and Ribonucleic Acid interference (RNAi). CRISPR/CAS9 vectors and RNAi are prepared and transfected into chondrocytes to knockdown the expression of the COL11a1 gene. Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR) is used to verify the successful knockdown of COL11a1 and a Western Blots is used for protein verification. Differential gene expression is determined by comparison of the COL11a1 knockdown chondrocyte model and the normal chondrocyte model. Additionally, activation of cell signaling pathways are determined by Luciferase assays. Down regulation of the COL11a1 gene is expected to enhance the expression of osteoarthritic factors. This data can be used to procure a treatment to combat these osteoarthritic occurrences.