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Full-Text Articles in Molecular Biology

Improving The Efficiency Of Crispr-Cas9: The Expression & Purification Of Cas9, Elaine Shults Jan 2016

Improving The Efficiency Of Crispr-Cas9: The Expression & Purification Of Cas9, Elaine Shults

Undergraduate Honors Theses

p53 is a transcription factor that plays a significant role in cell cycle regulation, senescence, and apoptosis. The p53 protein has been widely studied for its link to human cancer, and is estimated to be mutated in ~50% of all human cancers. This project focused on generating a p53-null A549 cell line using the genome editing technique CRISPR (Clustered Regularly-Interspaced Short Palindromic Repeats)-Cas9. CRISPR-Cas9 was employed to generate a simultaneous knock-in/knockout of mCherry and WTp53, respectively. The mCherry gene was inserted at the WTp53 endogenous translational start site (TSS), thereby rendering WTp53 inactive and mCherry “active” whenever WTp53 ...


A Computationally Designed Protein-Ligand Interaction Is Mechanically Robust, William John Van Patten Jan 2016

A Computationally Designed Protein-Ligand Interaction Is Mechanically Robust, William John Van Patten

Undergraduate Honors Theses

Protein-ligand interactions govern essential and ubiquitous biological processes such as immune response and gene regulation. Recently, the first computationally designed ligand-binding protein named DIG10.3 was developed by the Baker lab at the University of Washington. This artificially designed (rather than naturally evolved) ligand binding protein exhibited high affinity and selectivity to its target ligand, Digoxigenin (Dig). Such computationally designed ligand-binders offer promising capabilities in diagnostics and therapeutics for a wide range of diseases. By applying a mechanical force to a single DIG10.3::Digoxigenin interaction through atomic force microscope (AFM)-based single-molecule force spectroscopy (SMFS) we can extract unique ...


Application Of The Split Gfp System To Listeria Monocytogenes To Visualize The Virulence Factor Inlc, Dilara Batan Jan 2016

Application Of The Split Gfp System To Listeria Monocytogenes To Visualize The Virulence Factor Inlc, Dilara Batan

Undergraduate Honors Theses

Listeria monocytogenes (Lm) is an opportunistic pathogen that is able to survive in a range of environments and cell types, and therefore serves as an important model system for host-pathogen studies. Lm can enter mammalian cells and survive within these host cells by secreting a number of virulence proteins during these steps. In the literature, there are inconsistencies in the localizations of one of these effector proteins, InlC. In order to better understand the localizations of the Lm effector protein InlC in the live cell during infections, a split GFP approach is taken to fluorescently label the protein. This system ...