Modeling 3D Retinogenesis in Mouse Embryonic Stem Cells Following CRISPR-mediated Crx Knockdown

<p>An emerging technology known as three-dimensional (3D) tissue engineering has allowed scientists to mimic tissues found <em>in vivo.</em> Previous studies indicate that it is possible to differentiate dissociated mouse embryonic stem cells (mESCs) into 3D retinal tissues <em>in vitro</em><em> </em>(Bertacchi, 2015; Eiraku, 2012). The newly differentiated retinal tissues are said to encompass all of the major components found in retinal tissues. The generation of <em>in vitro</em> 3D tissues holds great potential in terms of patient-specific disease modeling. Although various diseases have been well-studied in animal models, there are limitations with regards to patient-specificity. The generation of animal models to study each specific mutation would be impractical and costly. With <em>in vitro</em> generation of tissues and organs, scientists have the capacity to generate disease models consisting of mutations specific to patients.</p> <p>There are various gene editing techniques available to generate and recreate disease-specific 3D disease models using stem cells. The newest gene editing technique known as Clustered Regularly Interspersed Short Palindromic Repeats (CRISPR) has allowed scientists to edit the genome with greater precision than other genome editing technologies like Zinc Finger Nucleases (ZFNs) or Transcription Activator-like Effector Nucleases (TALENs).</p> <p>In order to overcome current limitations on disease-specific model systems, we developed a protocol for differentiating mouse embryonic stem cells into 3D retinas <em>in vitro</em>. In addition, to confirm the utility of this 3D model for future disease-specific mutations, we utilized the CRISPR method as a means to knockdown the transcription factor CRX, which is known to regulate key developmental decisions during retinogenesis, and as such, mutations in CRX have resulted in various diseases such as Leber Congenital Amaurosis, Cone-Rod Dystrophy, and Retinitis Pigmentosa.</p>

Name thesis:degree_name

Master of Science

Level thesis:degree_level

Master's Thesis

Discipline thesis:degree_discipline

Biological Sciences

Year dc:date.available

2017

Author dc:creator

• Prasad, Pooja

Contributors dc:contributor

• Deepak Lamba, MBBS, PhD
• Mary B. Sevigny, PhD

• CRISPR
• Retina
• Organoid
• Crx
• Mouse Embryonic Stem Cells
• Knockdown
• Biology
• Developmental Biology
• Developmental Neuroscience
• Molecular and Cellular Neuroscience
• Molecular Genetics

Repository record dc:identifier

https://scholar.dominican.edu/masters-theses/279

OAI identifier oai:identifier

oai:scholar.dominican.edu:masters-theses-1287