Genetics worksheet

Assignment Problems: Lectures MN 0350 Genetics Fall 2020 1

0350 Genetics Fall 2020 Assignment Problems: Lectures MNO (1) The table below lists E.coli strains, including partial diploids, mutant for different components of the lac operon. For each determine if ß-galactosidase activity would be detected with and without lactose present (assume glucose is low; no includes very low levels). (OC is the same as O-)

Genotype No lactose With lactose

1 (1a) I+ P+ O+ Z+ Y+ 2 (1b) I+ P+ O+ Z+ Y- 3 (1c) I+ P+ OC Z- Y+ 4 (1d) I-d P+ O+ Z+ Y+ 5 (1e) IS P+ OC Z+ Y+ 6 (1f) I+ P+ O+ Z+ Y-/ I+ P- OC Z+ Y- 7 (1g) I+ P+ O+ Z+ Y+/ I+ P+ OC Z- Y+ 8 (1h) I- P+ O+ Z+ Y+/ I+ P- O+ Z+ Y+ 9 (1i) I+ P+ OC Z+ Y-/ I+ P+ O+ Z+ Y-

10 (1j) IS P+ O+ Z+ Y-/ I+ P+ OC Z+ Y-

11 (1k) I-d P+ O+ Z+ Y+/IS P+ O+ Z+ Y+ A: No/No B: Yes/No C: No/Yes D: Yes/Yes E: Could be Yes/Yes or could be No/No, it depends on how the mutant proteins interact

Assignment Problems: Lectures MN 0350 Genetics Fall 2020 2

(2) The expression of four different Drosophila genes, ant, hed, nog and bon, during embryonic development was investigated by generating antibodies against the gene products and then fixing and staining embryos. Interestingly it was found that they were all expressed in the same cells in early embryos, corresponding to the anterior half of the embryo. Below are diagrams of embryos showing where cells expressing the four different proteins are localized to in embryos, which consist of thousands of cells at this stage.

Ant, Hed, Nog and Bon protein expression in early Drosophila embryos, anterior (A) to the left. All four proteins are found in the same cells in the anterior half

The basic question to be addressed here is why is each protein expressed in the cells in the anterior region and why it is not expressed elsewhere. Although all four proteins are found in the same cells, subsequent analysis revealed that the regulatory mechanisms controlling transcription/translation of each gene/protein expression was different. The genomic region surrounding each gene was characterized and each gene was shown to have a single enhancer upstream that are required for the expression of each protein. Genetic analysis identified transcription factors and microRNAs that control expression of each gene and these are different for each gene:

Y for ant M for hed S and T for Nog S and H for Bon

i.e. Y, M, S, T and H could be transcription factors or microRNAs. Assume that these are the only factors regulating expression of the four genes. The diagram below shows the expression pattern of (i) each protein, (ii) Ant, Hed, Nog and Bon mRNA in wild- type embryos, (iii) each transcription factor or regulatory RNA (Y, M, S, T and H) and (iv) Ant, Hed, Nog and Bon mRNA in mutants of the genes that encode those transcription factors/regulatory RNAs.

Assignment Problems: Lectures MN 0350 Genetics Fall 2020 3

For each of the transcription factors or RNAs, Y, M, S, T, H, state whether each is most likely to encode a transcription factor - activator (active all the time or activity is regulated) or repressor, or a microRNA? 12 (2a) Y A Transcription factor: activator, always active B Transcription factor: repressor C Transcription factor: activator whose activity is regulated (could possibly be repressor when not activated) D microRNA 13 (2b) M A Transcription factor: activator, always active B Transcription factor: repressor C Transcription factor: activator whose activity is regulated (could possibly be repressor when not activated) D microRNA 14 (2c) S A Transcription factor: activator, always active B Transcription factor: repressor C Transcription factor: activator whose activity is regulated (could possibly be repressor when not activated) D microRNA 15 (2d) T A Transcription factor: activator, always active B Transcription factor: repressor C Transcription factor: activator whose activity is regulated (could possibly be repressor when not activated) D microRNA 16(2e) H A Transcription factor: activator, always active B Transcription factor: repressor C Transcription factor: activator whose activity is regulated (could possibly be repressor when not activated) D microRNA Inside a cell where would each of the factors be bound to perform its function? Below is an idealized diagram corresponding to each gene, with an enhancer, the coding region for the gene, transcription, splicing, translation ultimately leading to the expression of the protein. Also, depicted are potential sites where Y, M, S,. T and H would be situated to perform their function. (Note again this is an idealized diagram from each gene there are four genes, each with their own coding region and their own enhancer).

Assignment Problems: Lectures MN 0350 Genetics Fall 2020 4

Consider a cell in the anterior region of an embryo and identify where each of the factors Y, M, S, T and H would be bound. So, again, the diagram is generic, for each question below consider where the factor would be located to regulate the specific gene it controls (as indicated). 17 (2f) Y (for ant gene) A At the enhancer B At the promoter C On the mRNA D Not at locations A, B or C 18 (2g) M (for hed gene) A At the enhancer B At the promoter C On the mRNA D Would not be in the cell or at locations A, B, C 19 (2h) S (for nog or bon genes) A At the enhancer B At the promoter C On the mRNA D Not at locations A, B or C 20 (2i) T (for nog gene) A At the enhancer B At the promoter C On the mRNA D Not at locations A, B or C 21 (2j) H (for bon gene) A At the enhancer B At the promoter C On the mRNA D Not at locations A, B or C Consider a cell in the posterior region of an embryo identify where each of the factors Y, M, S, T and H would be situated (choose the best option).

Assignment Problems: Lectures MN 0350 Genetics Fall 2020 5

22 (2k) Y (for ant gene) A At the enhancer B At the promoter C On the mRNA D Not at locations A, B or C E Possibly at the enhancer or possibly not there or B or C 23 (2l) M (for hed gene) A At the enhancer B Definitely at the promoter C On the mRNA D Not at locations A, B or C E Possibly at the enhancer or possibly not there or B or C 24 (2m) S (just consider the nog gene, not the bon.) A At the enhancer B At the promoter C On the mRNA D Not at locations A, B or C E Possibly at the enhancer or possibly not there or B or C 25 (2n) T (for nog gene) A At the enhancer B At the promoter C On the mRNA D Not at locations A, B or C E Possibly at the enhancer or possibly not there or B or C 26 (2o) H (for bon gene) A At the enhancer B At the promoter C On the mRNA D Not at locations A, B or C E Possibly at the enhancer or possibly not there or B or C If the enhancer from each gene was hooked up to a GFP reporter gene (with promoter), would GFP show the same pattern of expression as each protein, i.e. would it only be expressed in the anterior, identically to each protein in embryos? 27 (2p) ant enhancer A Yes B No 28 (2q) hed enhancer A Yes B No

Assignment Problems: Lectures MN 0350 Genetics Fall 2020 6

29 (2r) nog enhancer A Yes B No 30 (2s) bon enhancer A Yes B No

Assignment Problems: Lectures MN 0350 Genetics Fall 2020 7

3 31 (3a) What is a transgenic organism? A: One which has a cloned DNA fragment inserted into its genome. This may be a gene from another species or one of its own genes, but if the latter it is in a different location to the endogenous gene. B: One which has any kind of artificial modification to its genome, such as a CRISPR/Cas induced mutation C: One in which any part of its genome has been moved to a different location. D: One in which part of one chromosome has been moved to a different region of the genome. 32 (3b) What enzyme is targeted by Roundup herbicide A: RNA polymerase B: Alanine synthase C: Adenine synthase D: EPSP synthase 33 (3c) What reagents are required to engineer plants to be resistant to roundup A: DNA plasmid containing a bacterial gene encoding the enzyme targeted by Roundup, microinjector, additional plasmid containing vir genes B: DNA plasmid containing a bacterial gene encoding the enzyme targeted by Roundup, Agrobacterium tumefaciens C: DNA plasmid containing a bacterial gene encoding the enzyme targeted by Roundup, Agrobacterium tumefaciens, additional plasmid containing vir genes D: DNA plasmid containing a bacterial gene encoding the enzyme targeted by Roundup, bacteriophage, additional plasmid containing vir genes 34 (3d) What sequences are essential in the plasmid containing the bacterial gene for its integration into the host genome A: Inverted repeats from the P-element B: attP and attB sequences from øC31 bacteriphage C: LB and RB sequences from the T plasmid D: None it will be integrated by host cell proteins that promote non-homologous recombination 35 (3e) Some sequences must be added to the bacterial gene for this experiment to work, which of the following is NOT essential A: Enhancer sequences B: Intron(s) C: Eukaryotic promoter sequences D: CTP sequence (you are going to have to look that one up yourself we didn’t talk about it in class) E: Eukaryotic transcriptional termination sequences

Assignment Problems: Lectures MN 0350 Genetics Fall 2020 8

36 (3f) Would it be possible to develop a Roundup-resistant plant without the use of genetic engineering just by using standard breeding techniques? A: Possibly, as some weeds are becoming resistant through natural mutations B: No, it would be impossible due to the fragile nature of crop plants C: Probably not, as such strains would have already evolved D: Plants are not infected by E. coli so it would be impossible to introduce a bacterial gene 37 (3h) Would it be possible to use CRISPR/Cas9 technology to generate a Roundup resistant plant? A: No, CRISPR/Cas9 is just used to make knockout mutations B: No, CRISPR/Cas9 can only be used to modify existing genes C: Yes, the endogenous gene could be replaced with a resistant version. D: Yes, a knockout of the gene would result in resistance