Crispr Gene Editing in Yeast: An Experimental Protocol for An Upper-Division Undergraduate Laboratory Course

Autor/inn/en	Sehgal, Nitasha; Sylves, M. Eileen; Sahoo, Ansuman; Chow, Jacky; Walker, Sarah E.; Cullen, Paul J.; Berry, James O.
Titel	Crispr Gene Editing in Yeast: An Experimental Protocol for An Upper-Division Undergraduate Laboratory Course
Quelle	In: Biochemistry and Molecular Biology Education, 46 (2018) 6, S.592-601 (10 Seiten)Infoseite zur Zeitschrift PDF als Volltext Verfügbarkeit
Zusatzinformation	ORCID (Berry, James O.)
Sprache	englisch
Dokumenttyp	gedruckt; online; Zeitschriftenaufsatz
ISSN	1470-8175
DOI	10.1002/bmb.21175
Schlagwörter	Genetics; Science Instruction; Science Laboratories; Scientific Research; Biotechnology; Undergraduate Study; College Science; Science Activities; Science Process Skills + Suchen Sie Ihr Suchwort? Humangenetik; Teaching of science; Science education; Natural sciences Lessons; Naturwissenschaftlicher Unterricht; Biotechnologie; Grundstudium
Abstract	Clustered regularly interspaced short palindromic repeats (CRISPR) are a revolutionary tool based on a bacterial acquired immune response system. CRISPR has gained widespread use for gene editing in a variety of organisms and is an increasingly valuable tool for basic genetic research, with far-reaching implications for medicine, agriculture, and industry. This lab is based on the premise that upper division undergraduate students enrolled in a Life Sciences curriculum must become familiar with cutting edge advances in biotechnology that have significant impact on society. Toward this goal, we developed a new hands-on laboratory exercise incorporating the use of CRISPR-Cas9 and homology directed repair (HDR) to edit two well-characterized genes in the budding yeast, "Saccharomyces cerevisiae". The two genes edited in this exercise, Adenine2 (ADE2) and Sterile12 (STE12) affect metabolic and developmental processes, respectively. Editing the premature stop codons in these genes results in clearly identifiable phenotypes that can be assessed by students in a standard laboratory course setting. Making use of this basic eukaryotic model organism facilitates a laboratory exercise that is inexpensive, simple to organize, set up, and present to students. This exercise enables undergraduate students to initiate and follow-up on all stages of the CRISPR gene editing process, from identification of guide RNAs, amplification of an appropriate HDR fragment, and analysis of mutant phenotypes. The organization of this protocol also allows for easy modification, providing additional options for editing any expressed genes within the yeast genome to produce new mutations, or recovery of existing mutants to wild type. (As Provided).
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Erfasst von	ERIC (Education Resources Information Center), Washington, DC
Update	2020/1/01