Emma Chesley Collaborators Anna Frederich Question 1 2 points Fall 2014 7 012 Problem Set 6420 Points total DUE WEDNESDAY 11 19 2014 AT 9 45AM ONLINE We should use microarray B The hybridizing sample in this case would be cDNA We should use microarray A The hybridizing sample in this case would be genomic DNA DNA microarray technology has many applications It can be used to determine how gene expression differs between cell types and it can also be used to genotype individuals You design two DNA microarrays Microarray A represents genomic DNA sequences Microarray B represents the coding sequence of each gene in the genome a Which microarray would you use to determine how gene expression differs between different kinds of human cells What would your hybridizing sample be in this case cDNA or genomic DNA b Which microarray would you use to genotype an individual What would your hybridizing sample be in this case cDNA or genomic DNA In lecture we mentioned that aside from a mutant hunt one can study the function of a gene by removing the gene and studying the phenotype of the organism when the gene is absent Two possible methods used to remove a gene are listed below a Describe each method in one sentence and in two sentences clarify how they differ from one another Gene knockout deletion 4 RNAi b Suppose you want to study an essential gene and you know that a homozygous deletion of this gene has an embryonic lethal phenotype Which one of the two techniques from part a would you use and why Question 2 6 5 points We should use the RNAi because it will cause the RNA to be degraded which is not lethal We can see what protein would no longer be encoded and not kill the embryo in the process Gene knockout Knockouts usually involve a loss of function We are able to nd the gene in the organism by looking at the loss of function phenotype because the gene is alway the opposite of the loss of function phenotype RNAi This is known as a knockdown RNA interference cause the RNA to be degraded so it cannot encode for a protein This is less effective than a knockout because rather than deleting a gene it destroys the RNA Region 7 Exon 2 Region 4 Intron 2 Region 5 Below is the schematic for a gene gene A that you decide to study using the gene deletion method You decide to design a construct to knock out gene A and to identify the cells in which the target gene is replaced or disrupted Intron 1 Exon 1 Region 2 Region 3 The construct needs to allow for a recombination event that replaces or disrupts your gene of interest and allows you to distinguish between a specific recombination event that disrupts your gene of interest and a random recombination event Include in your schematic the features Exon 3 Region 1 Region 6 c Below sketch the sequence of a construct that would allow you to do this that would allow you to do so d Explain how you would ensure that the recombination event happened e How would you distinguish between a random recombination event of your construct and a recombination event in which your gene was replaced or disrupted f Suppose you decide to knock this gene down using RNAi Which regions of the gene from the schematic in part a could you use in designing your RNA construct Which region could you not use Explain Positive selectable markers ensure that the recombinant event will happen To ensure that the recombination event happened we can insert the organisms with an antibiotic that would be toxic to all organisms whose genes did not recombine properly If the genes did not recombine properly then the positive and negative selectable markers would recombine thus causing the organism to die We could use the exons region 2 region 4 and region 6 We could not use introns region 3 and region 5 Question 3 5 points A single nucleotide polymorphism SNP is a DNA sequence variation occurring when a single base pair in the genome differs between members of a species or paired chromosomes in an individual By convention this base pair change is represented as one nucleotide A T C or G of the base pair a If the SNP does not change the amino acid sequence of a protein where could this SNP be Choose all that apply from coding sequence non coding sequence within the gene non coding sequence between the genes Explain your answer b Below is the pedigree of a family with a disease that shows 100 penetrance Coding sequence it would be a silent mutation and thus would not change the amino acid sequence of a protein Non coding sequence within the gene it would not change the amino acid sequence of the protein because it is not transcribed Non coding sequence between the genes it would not change the amino acid sequence of the protein because it is no translated i e it would be in an intron All the individuals that show the disease phenotype are shaded The two letters identify the two alleles of a SNP that is tightly linked to Gene B the gene that is associated with this disease For example G A indicates that on one of the chromosomes you would find a G a G C base pair and on other chromosome you would find an A an A T base pair Please note that unlike most rare diseases some of the individuals marrying into this family may be carriers Also assume that no recombination occurs between the SNP and gene B in the members of this family 2 1 C G C A 3 4 C C 5 G C 6 7 C C 8 9 G A 10 G C 11 C G 22 12 13 14 15 16 17 18 19 20 21 What allele of the SNP is tightly linked with the disease allele in individuals 1 2 4 5 and 7 23 C G C G The C allele use the letter D to represent the allele associated with the dominant phenotype and d to represent the allele associated with the recessive c List all possible genotypes at the B locus of following individuals in this pedigree Note Use the symbol XD Xd D or d where appropriate In each case phenotype d What is the probability of Individual 22 being affected e Assume that individual 21 is not a carrier of the disease allele What is the chance that individual 23 is a carrier of the disease allele Show your There is a 50 change that individual 22 will be affected Genotype at the B locus Individuals 2 5 9 DD Dd Dd work There is no change because we know that individual 20 is not a carrier and we are told that individual 21 is not a carrier and so they can not have an offspring that is a carrier of the disease allele Question 4 3 5 points In the following pedigree individuals that have …
View Full Document
Unlocking...