Title

Using CRISPR-CAS9 to Investigate Whether Lysophosphatidic Acid Receptor 4 (LPAR4) Has a Role In Chick Retinal Ganglion Cell Axon Guidance

School Name

South Carolina Governor's School for Science & Mathematics

Grade Level

12th Grade

Presentation Topic

Cell and Molecular Biology

Presentation Type

Mentored

Abstract

During the development of a chick embryo, specific neurons in the developing eye, called retinal ganglion cells, send out axons that are guided to their terminate centers in the brain with the help of growth cones. Growth cones are the leading ends of axons and are guided by attractive or repulsive environmental cues. Previous studies have shown that lysophosphatidic acid (LPA) is a repelling axon guidance molecule that acts on the growth cones in a dose-dependent fashion. When LPA comes into contact with these retinal growth cones, they collapse and regrow in a different direction. LPA binds to six known G-coupled receptors (LPAR 1-6), but LPAR5 is not expressed in the retina. Previous research has shown that mice with mutated LPARs 1-3 had behaved normally hen tested individually, and, therefore it can be hypothesized that these receptors have no role in retinal axon guidance. The primary aim of this project is to determine whether LPAR4 might be the receptor that is responsible for causing growth cone collapse, in vitro. The CRISPR technique will be used to mutate LPAR4 and determine its effect on retinal ganglion cells. During the research, the negative control in the PCR showed a band that was similar to LPAR4. Various modifications were made to the protocol, and the primers were found to be the cause of the problem. Finally, a 180bp band was seen with new primers. Due to time constriction, CRISPR could not be accomplished. Future research will aim at making a mutated LPAR4.

Location

Furman Hall 107

Start Date

3-28-2020 8:30 AM

Presentation Format

Oral Only

Group Project

No

COinS
 
Mar 28th, 8:30 AM

Using CRISPR-CAS9 to Investigate Whether Lysophosphatidic Acid Receptor 4 (LPAR4) Has a Role In Chick Retinal Ganglion Cell Axon Guidance

Furman Hall 107

During the development of a chick embryo, specific neurons in the developing eye, called retinal ganglion cells, send out axons that are guided to their terminate centers in the brain with the help of growth cones. Growth cones are the leading ends of axons and are guided by attractive or repulsive environmental cues. Previous studies have shown that lysophosphatidic acid (LPA) is a repelling axon guidance molecule that acts on the growth cones in a dose-dependent fashion. When LPA comes into contact with these retinal growth cones, they collapse and regrow in a different direction. LPA binds to six known G-coupled receptors (LPAR 1-6), but LPAR5 is not expressed in the retina. Previous research has shown that mice with mutated LPARs 1-3 had behaved normally hen tested individually, and, therefore it can be hypothesized that these receptors have no role in retinal axon guidance. The primary aim of this project is to determine whether LPAR4 might be the receptor that is responsible for causing growth cone collapse, in vitro. The CRISPR technique will be used to mutate LPAR4 and determine its effect on retinal ganglion cells. During the research, the negative control in the PCR showed a band that was similar to LPAR4. Various modifications were made to the protocol, and the primers were found to be the cause of the problem. Finally, a 180bp band was seen with new primers. Due to time constriction, CRISPR could not be accomplished. Future research will aim at making a mutated LPAR4.