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Using Plasmids and the Transformation of E.Coli

The ampicillin resistance gene is included in the pRED, pGLO and pUC plasmids. Ampicillin resistance is included in these plasmids as a selection gene, also called a selection marker.

1) Explain how selection genes work

2) Why is it necessary to have a selection gene in plasmids that are used for genetic engineering and transformation?

3) Which plasmid - pRED, pGLO or pUC is the best/most efficient at transforming E. coli in laboratory experiments?
Note: pGLO plasmid is for ampicillin resistance and green fluorescent protein. Each gene has its own promoter, the amp promoter is constitutive, but the GFP is under an inducible promoter. The inducer is the sugar arabinose.
pRED plasmid is for ampicillin resistance and red fluorescent protein. Both genes are under constitutive promoters.
pUC promoter is for ampicillin resistance only

4) pUC plasmid is for ampicillin resistance. What does the gene product of the ampicillin resistance gene do? How does it work? How does it protect E. coli from the ampicillin antibiotic?

Solution Preview

The ampicillin resistance gene is included in the pRED, pGLO and pUC plasmids. Ampicillin resistance is included in these plasmids as a selection gene, also called a selection marker.

1) Explain how selection genes work

Genes that are under a selectable marker such as ampicillin resistance are called selection genes because they can be screen using plates that are ampicillin resistance. In the case of the pRED, pGLO, and pUC plasmids that have ampicillin resistance gene, they can be selected by tracking the growth of E. coli that are ampicillin resistance.
Transformation is the process of changing the E. coli genotype by introducing a foreign gene ( antibiotic resistance gene) on the plasmid into the E. coli cell. Once the foreign gene (antibiotic resistanc gene) on the plasmid enter the cell, it is expressed and change both the genotype and phenotype of the E. coli cell. To select for cell that expressed the antibiotic resistance gene, one would need plate that have the antibiotic ampicillin on them and plate for the transformant. If the transformant take up the plasmid with the antibiotic selection gene, then they will be resistant to the antibiotic on the plate and grow on the plate. If the transformants did not take up the antibiotic gene, they did not express the selection gene (antibiotic resistance gene) and will not grow on the antibiotic containing plate. The entry of the plasmid with the selective marker gene (antibiotic resistance) will change the genotype ...

Solution Summary

Genetic transformation is the process of altering the genotype and phenotype of a cell by the introduction of foreign DNA from external source. This solution explains gene selection, how to pick plasmids for different purposes in testing and resistance genes in just under 1000 words.

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