An undergraduate laboratory class using CRISPR/Cas9 technology to mutate drosophila genes

Abstract

CRISPR/Cas9 genome editing technology is used in the manipulation of genome sequences and gene expression. Due to the ease and rapidity with which genes can be mutated using CRISPR/Cas9, we sought to determine if a single-semester undergraduate class could be successfully taught, wherein students isolate mutants for specific genes using CRISPR/Cas9. Six students were each assigned a single Drosophila gene, for which no mutants currently exist. Each student designed and created plasmids to encode single guide RNAs that target their selected gene; injected the plasmids into Cas9-expressing embryos, in order to delete the selected gene; carried out a two-generation cross to test for germline transmission of a mutated allele and generate a stable stock of the mutant; and characterized the mutant alleles by PCR and sequencing. Three genes out of six were successfully mutated. Pre- and post- survey evaluations of the students in the class revealed that student attitudes towards their research competencies increased, although the changes were not statistically significant. We conclude that it is feasible to develop a laboratory genome editing class, to provide effective laboratory training to undergraduate students, and to generate mutant lines for use by the broader scientific community.

CRISPR-Cas Technology In and Out of the Classroom

“…In some cases, these experiences can be used to generate both laboratory reagents and data as well as data pertaining to student outcomes resulting from the implementation of CUREs. [21][22][23][24][25][26] Given student interest and the increased need to communicate cutting-edge research to undergraduate students, colleges and universities have begun instituting CRISPR-Casrelated CUREs with great success. Part of the reason why CUREs and other forms of course-based research that utilize CRISPR-Cas technologies are successful is due to the current curricula in molecular biology-based laboratory courses at diverse academic institutions, particularly fouryear colleges and universities.…”

“…Although data collection on the outcomes of CRISPR-Cas-based CUREs is ongoing, instructors report increased student learning and engagement after implementing CRISPR-Cas-based CUREs using various metrics. [21][22][23][24][25][26] Academic institutions have only recently started using CRISPR-Cas-based CUREs in their coursework; with increased interest in CRISPR-Cas-based CUREs, more rigorous studies on student achievement will undoubtedly help inform future activities at more institutions.…”

“…Importantly, although Dr. Richard Cripps only just introduced this course in 2015, he has since published his laboratorybased course model, including the data generated by his students during the course. 24 Some CUREs allow faculty researchers to generate valuable tools and/or reagents that can be used in independent research projects for students within the faculty member's laboratory. For instance, Dr. Kevin Militello at the State University of New York, Geneseo recently published a paper illustrating the power of his laboratorybased course model wherein students examined the CRISPR loci of uncharacterized E. coli strains.…”

“…[22][23][24]28,30,31 Because CUREs that utilize CRISPR-Cas technology are relatively new, there is limited information about how these academic gains translate into student career choices or persistence in scientific (and particularly those involving genetic engineering) fields. However, as the availability of CRISPR-Cas-based CUREs increases, more information regarding the benefits to students will likely become available.…”

Absence of the Drosophila jump muscle actin Act79B is compensated by up‐regulation of Act88F

“…Recently, Adame et al (2016) demonstrated that Drosophila Act79B null mutants appeared to jump normally, despite lacking the predominant actin found in the jump muscle (also called the tergal depressor of the trochanter, TDT). However, the sample size was small; therefore, in this study we investigated further the role of Act78B and other actin isoforms in the jump muscle of Drosophila.…”