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  • About
  • The Global ETD Search service is a free service for researchers to find electronic theses and dissertations. This service is provided by the Networked Digital Library of Theses and Dissertations.
    Our metadata is collected from universities around the world. If you manage a university/consortium/country archive and want to be added, details can be found on the NDLTD website.
1

Investigating cellular functions of the SMARCAD1 gene in human MPNST cells by CRISPR-Cas13d knockdown

Han Han (12442215) 22 April 2022 (has links)
<p>  </p> <p>Malignant Peripheral Nerve Sheath Tumor (MPNST) is a form of soft tissue sarcoma arising from peripheral nerve sheath cells. Currently, there is no clinically available targeted therapy because the targetable essential driver genes in this tumor are largely unknown. SMARCAD1 (SWI/SNF-related, matrix-associated actin-dependent regulator of chromatin, subfamily A, containing DEAD/H box 1) has been identified as a new tumor suppressor of MPNSTs in zebrafish. Several studies have also linked <em>SMARCAD1</em> with cancer development together. However, the cellular roles of <em>SMARCAD1</em> in human MPNST cells remain unclear. To investigate DNA damage repair functions of SMARCAD1 in human MPNST, we created a doxycycline-inducible Schwannoma cell line by CRISPR-Cas13d, a newly developed mRNA knockdown method. I verified efficiently SMARCAD1 knockdown cell line by western blot. In addition, knockdown of SMARCAD1 inhibits Schwannoma cell proliferation and anchorage-independent growth. It is reported that SMARCAD1 is involved in DNA damage repair mechanisms. I confirmed that loss of SMARCAD1 expression compromises DNA damage repairing function in Schwannoma cells. This result was also verified in two zebrafish <em>smarcad1</em> mutants. In summary, I utilized a novel gene knockdown approach to generate a SMARCAD1 Schwannoma cell line and validated its function in DNA damage repair. This study might provide information for developing a new treatment option for MPNSTs.</p>
2

Harnessing a novel compact CRISPR-Cas13b for SARS-CoV-2 diagnostics

Wang, Qiaochu 04 1900 (has links)
The outbreak of infectious diseases across the world results in huge disasters for public health. Rapid and effective diagnostic methods are crucial for disease identification and transmission control. Since first identified in late 2019, the pandemic of COVID-19 caused by the SARS-CoV-2 virus resulted in unprecedented catastrophe globally. To control the further spread of COVID-19, there is an urgent need for rapid, accurate, cost-effective, and efficient diagnostics. Recently, many CRISPR-based diagnostics have been developed by coupling isothermal amplification methods with Cas proteinmediated nucleic acid detection. Compared with conventional methods like RT-qPCR, CRISPR-based assays are more cost-effective and efficient without sacrificing sensitivity and specificity. Here, I developed a Cas13-based assay for SARS-CoV-2 detection with a novel compact Cas13b protein. In this assay, the Cas13 detection is combined with RT-LAMP, achieving the detection of viral RNA as low as 4 copies/μl. By utilizing a simple LED-based visualizer (P51™) instead of a plate reader, the detection result can be visualized directly without using sophisticated instruments. The compact Cas13b-mediated viral detection together with P51™-based visualization enable rapid, sensitive, and portable diagnostics for SARS-CoV-2, showing great potential in application to point-of-care testing.

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