Genetics Lab Final Exam Study Guide

Genetics Lab Final Exam Study Guide

• The exam will have 30-35 questions, a mix of multiple choice, true/false, and written response.
• It will cover both theoretical and practical applications.
• Students should be prepared to explain homology and its role in model organism development.
• Identifying homologous genes from different organisms given a gene sequence is important.
• Students need to identify sgRNAs with adjacent PAM domains.
• Understanding in-vitro CRISPR experiment results and identifying potential errors is crucial.
• Interpreting gel results from experiments is a key skill.
• Implementing correct micropipetting techniques is vital.

Module 1: Mosquitoes and Reverse Genetics

• Mosquitoes (order Diptera) have a strong water dependence.
• Aedes aegypti, a common vector for diseases, is a model organism studied in this course.
• The genes IR68a and OBP71 are potentially involved in mosquito water sensing.
• Homology in genes allows inferences about function across different organisms.
• Reverse genetics involves creating a mutant genotype and studying its phenotype.
• Forward genetics is the traditional approach of observing a mutant phenotype to find the causative gene.
• A 2x2 factorial design is used for reverse genetics experiments with a genetic background (e.g., wild-type or mutant) and a treatment variable.
• Positive controls ensure the experiment functions correctly, while negative controls show what "no treatment" looks like.

Module 2: Pipetting Techniques

• Students need to understand pipette usage, specifically the calibrated volumes and how to read the volume dial.
• Understanding volume relationships is crucial.

Module 3: Gene Families and BLAST

• Gene families are formed through gene duplication within a genome creating paralogs.
• Speciation creates orthologs.
• Homologous genes can be compared using BLAST tools.
• Students should be familiar with different BLAST types, how they function, and interpret outputs like e-value, query cover and percent identity.

Module 4: Gene Structure and DNA Extraction

• Exons code for proteins and introns do not.
• Exons, introns, and UTRs (untranslated regions) are transcribed into pre-mRNA.
• Pre-mRNA undergoes processing to produce mRNA with only exons.
• mRNA is translated into proteins.
• Gel electrophoresis is used to visualize DNA.
• Extraction techniques like homogenization, cell lysis, DNA binding, washing, and elution are used to purify DNA samples.
• Students should study the role of reagents in DNA extraction.

Module 5: Genotyping and PCR

• Genotyping typically starts by amplifying a region.
• PCR, or polymerase chain reaction, is used to amplify target sequences in repeated cycles.
• Each cycle doubles the target DNA amount.
• The PCR process includes denaturation, annealing, and elongation steps.

Module 6, 7: Gel Electrophoresis and Sequencing

• Gel electrophoresis separates DNA fragments by size.
• DNA is negatively charged and moves toward the positive electrode.
• Smaller fragments travel farther in the gel.
• Gel electrophoresis can be used to detect small indels (Insertions and/or Deletions).
• Techniques like fragment analysis give a digital output instead of a gel.
• Restriction digests can be used to identify mutations that disrupt restriction sites for identification.
• Cycle sequencing and Sanger sequencing are used to determine DNA sequences.

Module 8: CRISPR-Cas9

• CRISPR-Cas9 is a genome editing technology targeting DNA sequences using an RNA-guided nucleases.
• CRISPR-Cas9 creates small insertions or deletions (indels) in DNA.
• Methods used and their relationship to knock-out mutants and frameshift mutations.
• Frame-shift mutations disrupt the reading frame of codons.
• Understanding the use of PAM (protospacer adjacent motif) regions, and their relationship to the target sequence.

Review: Sanger Sequencing and Gel Electrophoresis

• Students should be able to interpret a Sanger sequencing gel electrophoresis.
• Understand the concept of polymorphism.