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- To produce transgenic plants, plant tissue is exposed to Agrobacteriumtumefaciens and then grown in media containing kanamycin, carbenicillin, and plant growth hormones. Explain the purpose behind each of these three agents. What would happen if you left out the kanamycin?Besides using A. tumefaciencs in the generation of transgenic plants, elaborate on the THREE (3) methods to introduce genc of interest into a plant cell.Outline a PCR-based approach to screen for colonies transformed with recombinant expression plasmid that can be used for protein expression.
- What are site-recombinases? Describe in detail how cre- recombinase can be used to decipher the roles of specific genes and proteins in complex multicellular organisms? Explain how the cre-lox system can be used to examine the role of a particular gene in a specific type of tissue?There are very different rules governing genome edited crops in the European Union and in the United States of America, according to Cohen. Explain why CRISPR-edited crops could be considered to be a type of GMO (per the EU) and why they could be considered equivalent to naturally occurring mutations (per the USA). In your explanation be sure to (1) discuss how CRISPR constructs are currently introduced into plants and (2) how all evidence of the CRISPR apparatus/process can be eliminated from the genome, leaving only the edited sequence(s).Agrobacterium-mediated plant transformation is a method used to create transgenic plants. Agrobacterium tumefaciens, a soil bacterium works exceptionally well as a natural genetic engineer. Describe briefly the creation of a transgenic plant using A. tumefaciens with illustration.
- What is homologous recombination and where/when does it typically take place in an organism. Homologous recombination is also used experimentally during gene targeting. If you wanted to remove the function of the mouse gene fibroblast growth factor 4 (FGF4) using gene targeting by homologous recombination, what are the different aspects of a targeting vector you would need to make?What are the following features that are required facilitate cloning into a vector.Briefly and in simple terms, describe how CRISPR could be utilized in treating genetic conditions such as Batten disease.
- Describe the rationale behind the electrophoretic mobility shift assay.There is a hypothetical gene related to the nervous system of Drosophila. Describe all the methods, steps, and key substances you need to obtain to use the following techniques in experimental design to study the gene: - In situ hybridization (to find the mRNA) - Immunohistochemistry (to find the protein) - CRISPR-Cas9 (for loss of function) - Expression vector (for gain of function)Besides cloning vectors, there are also expression vectors. What is the main difference between these two vectors? Give two (2) cxamples of expression vectors commonly used for prokaryotic cells.