"Genetics BIO243" 4 posts Sort by created date Sort by defined ordering View as a grid View as a list

CRISPR/Cas9 in yeast: a multi-week laboratory exercise for undergraduate students

Providing undergraduate life-science students with a course-based research experience that utilizes cutting-edge technology, is tractable for students, and is manageable as an instructor is a challenge. Here, I describe a multi-week lesson plan for a laboratory-based course with the goal of editing the genome of budding yeast, Saccharomyces cerevisiae. Students apply knowledge regarding advanced topics such as: CRISPR/Cas9 gene editing, DNA repair, genetics, and cloning. The lesson requires students to master skills such as bioinformatics analysis, restriction enzyme digestion, ligation, basic microbiology skills, polymerase chain reaction, and plasmid purification. Instructors are led through the technical aspects of the protocols, as well as the teaching philosophy involved throughout the laboratory experience. As it stands, the laboratory lesson is appropriate for 6-8 weeks of an upper-level undergraduate laboratory course, but may be adapted for shorter stints and students with less experience. Students complete the lesson with a more realistic idea of life science research and report significant learning gains. I anticipate this lesson to provide instructors and students in undergraduate programs with a hands-on, discovery-based learning experience that allows students to cultivate skills essential for success in the life sciences.

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Tackling "Big Data" with Biology Undergrads: A Simple RNA-seq Data Analysis Tutorial Using Galaxy

Analyzing high-throughput DNA sequence data is a fundamental skill in modern biology. However, real and perceived barriers such as massive file sizes, substantial computational requirements, and lack of instructor background knowledge can discourage faculty from incorporating high-throughput sequence data into their courses. We developed a straightforward and detailed tutorial that guides students through the analysis of RNA sequencing (RNA-seq) data using Galaxy, a public web-based bioinformatics platform. The tutorial stretches over three laboratory periods (~8 hours) and is appropriate for undergraduate molecular biology and genetics courses. Sequence files are imported into a student's Galaxy user account directly from the National Center for Biotechnology Information Sequence Read Archive (NCBI SRA), eliminating the need for on-site file storage. Using Galaxy's graphical user interface and a defined set of analysis tools, students perform sequence quality assessment and trimming, map individual sequence reads to a genome, generate a counts table, and carry out differential gene expression analysis. All of these steps are performed "in the cloud," using offsite computational infrastructure. The provided tutorial utilizes RNA-seq data from a published study focused on nematode infection of Arabidopsis thaliana. Based on their analysis of the data, students are challenged to develop new hypotheses about how plants respond to nematode parasitism. However, the workflow is flexible and can accommodate alternative data sets from NCBI SRA or the instructor. Overall, this resource provides a simple introduction to the analysis of "big data" in the undergraduate classroom, with limited prior background and infrastructure required for successful implementation.

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DNA Detective: Genotype to Phenotype. A Bioinformatics Workshop for Middle School to College.

Advances in high-throughput techniques have resulted in a rising demand for scientists with basic bioinformatics skills as well as workshops and curricula that teach students bioinformatics concepts. DNA Detective is a workshop we designed to introduce students to big data and bioinformatics using CyVerse and the Dolan DNA Learning Center's online DNA Subway platform. DNA Subway is a user-friendly workspace for genome analysis and uses the metaphor of a network of subway lines to familiarize users with the steps involved in annotating and comparing DNA sequences. For DNA Detective, we use the DNA Subway Red Line to guide students through analyzing a "mystery" DNA sequence to distinguish its gene structure and name. During the workshop, students are assigned a unique Arabidopsis thaliana DNA sequence. Students "travel" the Red Line to computationally find and remove sequence repeats, use gene prediction software to identify structural elements of the sequence, search databases of known genes to determine the identity of their mystery sequence, and synthesize these results into a model of their gene. Next, students use The Arabidopsis Information Resource (TAIR) to identify their gene's function so they can hypothesize what a mutant plant lacking that gene might look like (its phenotype). Then, from a group of plants in the room, students select the plant they think is most likely defective for their gene. Through this workshop, students are acquainted to the flow of genetic information from genotype to phenotype and tackle complex genomics analyses in hopes of inspiring and empowering them towards continued science education.

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Using computational molecular modeling software to demonstrate how DNA mutations cause phenotypes

Students require a deep understanding of the central dogma before they can understand complex topics such as evolution and biochemical disorders. However, getting undergraduate biology students to apply higher-order thinking skills to the central dogma is a challenge. Students remember and regurgitate the molecular details of transcription and translation but if asked to apply these details, such as how a DNA mutation might affect phenotype, it becomes clear that most students do not deeply understand the central dogma. This lesson is a five-week series of laboratory activities designed to help students transition from applying lower order thinking skills to the central dogma to applying higher-order thinking skills. Over five weeks, students explore the phenotype of Arabidopsis asymmetric leaves 1 (as1) and as2 mutants. Students isolate DNA from wild-type and mutant plants and determine the sequence of the AS1 and AS2 alleles. Students use the DNA sequence data to determine the mutant protein amino acid sequences. They submit the mutant and wild-type protein sequences to a free online server and obtain three-dimensional (3-D) models of the wild-type and mutant proteins. They use free software to analyze and compare the 3-D models to determine the structural differences between the wild-type and mutant proteins. These computer-generated models can be 3-D printed allowing students to better visualize the protein structure. The overall goal is to use student-centered laboratory activities to demonstrate the relationship between DNA sequence, protein structure/function, and phenotype.

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