國立虎尾科技大學 |

Elucidation of miRNA Function Using the CRISPR-Cas9 System.

Record Type:	Language materials, manuscript : Monograph/item
Title/Author:	Elucidation of miRNA Function Using the CRISPR-Cas9 System./
Author:	Kurata, Jessica Setsuko.
Description:	1 online resource (146 pages)
Notes:	Source: Dissertation Abstracts International, Volume: 79-12(E), Section: B.
Contained By:	Dissertation Abstracts International79-12B(E).
Subject:	Biology. -
Online resource:	click for full text (PQDT)
ISBN:	9780438267169

Elucidation of miRNA Function Using the CRISPR-Cas9 System.
Kurata, Jessica Setsuko.

Elucidation of miRNA Function Using the CRISPR-Cas9 System. - 1 online resource (146 pages)

Source: Dissertation Abstracts International, Volume: 79-12(E), Section: B.

Thesis (Ph.D.)--City of Hope's Irell & Manella Graduate School of Biomedical Sciences, 2018.

Includes bibliographical references

MicroRNAs (miRNAs) play an important role in the regulation of gene expression and are often dysregulated in disease. The recent development of the CRISPRCas9 gene-editing system, composed of the Cas9 nuclease in complex with a single guide RNA (sgRNA), allows researchers to direct DNA cleavage and mutation at a predetermined site. I used CRISPR-Cas9 to investigate miRNA function by both conducting genome-scale knockout screens and knocking out a single miRNA of interest.

Electronic reproduction.
Ann Arbor, Mich. :
ProQuest,
2018

Mode of access: World Wide Web

ISBN: 9780438267169Subjects--Topical Terms:

599573
Biology.
Index Terms--Genre/Form:

554714
Electronic books.

Elucidation of miRNA Function Using the CRISPR-Cas9 System.
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245 1 0 $a Elucidation of miRNA Function Using the CRISPR-Cas9 System.
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300 $a 1 online resource (146 pages)
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500 $a Source: Dissertation Abstracts International, Volume: 79-12(E), Section: B.
500 $a Includes supplementary digital materials.
500 $a Advisers: Ren-Jang Lin; Shizhen Wang.
502 $a Thesis (Ph.D.)--City of Hope's Irell & Manella Graduate School of Biomedical Sciences, 2018.
504 $a Includes bibliographical references
520 $a MicroRNAs (miRNAs) play an important role in the regulation of gene expression and are often dysregulated in disease. The recent development of the CRISPRCas9 gene-editing system, composed of the Cas9 nuclease in complex with a single guide RNA (sgRNA), allows researchers to direct DNA cleavage and mutation at a predetermined site. I used CRISPR-Cas9 to investigate miRNA function by both conducting genome-scale knockout screens and knocking out a single miRNA of interest.
520 $a To determine the functional role of miRNAs in cancer, we designed and constructed the LX-miR library of 7,382 sgRNAs to target 85% of the 1,881 annotated human miRNA stem-loops. I then examined the role of miRNAs in HeLa cell fitness by monitoring the change in frequency of each sgRNA over time. I identified 44 pro-proliferative miRNAs from two replicate experiments, including miR-31, a known cervical cancer overexpressing miRNA that enhances HeLa cell proliferation. I also examined the role of miRNAs in NCI-N87 gastric cancer cells and identified 10 pro-fitness and 10 anti-fitness miRNAs. In both screens, many of the pro-fitness miRNAs identified are overexpressed in tumor samples.
520 $a One of the identified HeLa pro-fitness miRNAs, miR-151a, was further investigated by using CRISPR-Cas9 to create HeLa clonal cell lines with decreased or complete loss of miR-151a expression. The knockout or knockdown of miR-151a was found to decrease cell fitness, in agreement with what was seen in the LX-miR screen. I also found there was an increase in the fraction of cells in the G1-phase of the cell cycle after miR-151a knockdown. This increase corresponded to an increase in p53 and p21 protein levels. I found miR-151a-3p is able to directly suppress p53 expression and that the increase in p21 detected is partially due to the increase in p53. By analyzing TCGA's database of paired tumor and normal samples, I discovered increased miR-151a expression is common in virtually all tumors where data are available. There was also an inverse correlation between miR-151a and p21 mRNA expression in approximately half of cancers for which there is miRNA sequencing information in TCGA. In summary, I present a CRISPR miRNA-targeted screen which identified both known and novel fitness-associated miRNAs in two cancer cell lines. I verified the importance of miR-151a for the fitness of HeLa cells and found miR-151a regulates the cell cycle through p53 and p21. Results from this study identify a previously unappreciated role of miR-151a in the regulation of the cell cycle.
533 $a Electronic reproduction. $b Ann Arbor, Mich. : $c ProQuest, $d 2018
538 $a Mode of access: World Wide Web
650 4 $a Biology. $3 599573
655 7 $a Electronic books. $2 local $3 554714
690 $a 0306
710 2 $a ProQuest Information and Learning Co. $3 1178819
710 2 $a City of Hope's Irell & Manella Graduate School of Biomedical Sciences. $b Graduate School of Biological Sciences. $3 1195349
773 0 $t Dissertation Abstracts International $g 79-12B(E).
856 4 0 $u http://pqdd.sinica.edu.tw/twdaoapp/servlet/advanced?query=10840660 $z click for full text (PQDT)