These are homework exercises to accompany Nickle and Barrette-Ng's "Online Open Genetics" TextMap. Genetics is the scientific study of heredity and the variation of inherited characteristics. It includes the study of genes, themselves, how they function, interact, and produce the visible and measurable characteristics we see in individuals and populations of species as they change from one generation to the next, over time, and in different environments.
11.1 What are the advantages of high-throughput –omics techniques compared to studying a single gene or protein at a time? What are the disadvantages
11.2 What would the chromatogram from a capillary sequencer look like if you accidentally added only template, primers, polymerase, and fluorescent terminators to the sequencing reaction?
11.3 What are the advantages and disadvantages of clone-by-clone vs. whole genome shotgun sequencing?
11.4 How could you use DNA sequencing to identify new species of marine microorganisms?
11.5 Explain how you could use a microarray to identify wheat genes that have altered expression during drought?
11.6 A microarray identified 100 genes whose transcripts are abundant in tumors, but absent in normal tissues. Do any or all of these transcripts cause cancer? Explain your answer.
11.7 How could you ensure that each spot printed on a microarray contains DNA for only one gene?
11.8 What would the spots look like on a microarray after hybridization, if each spot contained a random mixture of genes?
11.9 What would the spots look like if the hybridization of green and red labeled DNA was done at low stringency?
11.1 -omics techniques allow you to examine thousands of genes or protein at the same time. On a per-gene basis, this is more cost effective than examining a single gene at a time. However, the total cost of –omics technologies can be quite high (especially the instrumentation). Besides being more efficient, by examining many genes in parallel, you may identify patterns or combinations of genes that work together.
11.2 If only terminators (but no regular dNTPs) were added to the reaction, the reaction would always terminate at the first base added after the primer, and chromatogram would be essentially flat lines (no peaks) for all but the first position, which would show a Brobdingnagian peak.
11.3 Clone-by-clone sequencing requires less sequencing than WGS, since the objective is to minimize sequencing of redundant fragments. However, clone-by-clone is ultimately less efficient, since it takes so long to determine the minimum number of clones to sequence, through physical mapping. WGS is faster and more cost-effective, but is more prone to result in gaps in the sequence than is clone-by-clone sequencing.
11.4 You could extract raw, naked DNA from seawater in various different places in the world and then sequence all of this DNA. Next, use computer comparisons to identify DNA that did not belong to any known species. This is an example of meta-genomics, and is already being done by some scientists. Remember, having the sequence is not the same as having the organism or understanding the sequence.
11.5 One way is to extract RNA from (1) a drought-stressed and (2) a well-watered wheat plant. Reverse transcribe (make a DNA copy) the RNA samples independently and label each with a different fluorescent dye, then hybridize the two sample simultaneously to a microarray of wheat genes. If the drought-stressed sample was labeled one colour, say red, and the watered plant another colour, say green, then those gene transcripts that increase in abundance specifically under drought will be reder, while those that decreased would be greener, relatively speaking.
11.6 All that microarrays and similar types of analyses demonstrate (whether high-throughput or not) is a correlation between transcript presence/abundance and a phenotype. This does not mean that the genes cause cancer, although some of them could. The alternate probable explanation is that the gene’s change in expression is a response to being cancerous.
11.7 PCR amplify DNA from only a single bacterial clone containing single insert of part of a gene.
11.8 Spots with a random mix would be approximately equally green and red, which appears yellow or brown in the microarray analysis equipment.
11.9 At low stringency there would be excessive non-specific hybridization so many (all?) the spots would be approximately equally green and red, which appears yellow or brown in the microarray analysis equipment.
Dr. Todd Nickle and Isabelle Barrette-Ng (Mount Royal University) The content on this page is licensed under CC SA 3.0 licensing guidelines.