Question

Order the steps that would be used in a laboratory to engineer a bacterium that could express the human gene coding for factor VIII. Not all steps will be placed.Isolate the human gene that produces factor VIII-Generate cDNA of the factor VIII gene using reverse transcriptase- Transform the vector into E. coli bacterium-The factor VIII protein is isoloated from human tissues-Insert the factor VIII cDNA into a bacterial vector near a promoter site-Human DNA is introduced into the bacterial cell using direct injection by a small needle-Isolate the mRNA of the factor VIII geneE.coli expresses factor VIII

Answer 1

Order of steps in the laboratory to engineer a bacterium that could express the human gene coding for factor VIII:

1. Isolate the human gene that produces factor VIII
2. Generate cDNA of the factor VIII gene using reverse transcriptase
3. Insert the factor VIII cDNA into a bacterial vector near a promoter site
4. Transform the vector into E. coli bacterium
5. E.coli expresses factor VIII
6. The factor VIII protein is isolated from human tissues

Step-by-step explanation:

To engineer a bacterium to express the human gene for Factor VIII, a process known as recombinant DNA technology is employed. The following steps outline the procedure typically used in a laboratory setting:

1. Isolate the mRNA of the factor VIII gene from human cells.
2. Generate cDNA of the factor VIII gene using reverse transcriptase. This cDNA represents the coding sequence of the gene without introns, allowing bacterial expression.
3. Insert the factor VIII cDNA into a bacterial vector near a promoter site to ensure that the gene can be expressed by the bacteria.
4. Transform the recombinant vector into E. coli bacterium. This step involves introducing the vector containing the human gene into the bacterial cells.
5. Select for the successfully transformed bacteria that have incorporated the recombinant DNA.
6. Grow the selected bacteria, which express human factor VIII, and then isolate the protein produced.

These steps lead to the production of the factor VIII protein in bacterial cells, which can then be purified and utilized for various medical applications.

Answer 2

Isolate the mRNA of the factor VIII.

Generate cDNA of the factor VIII gene using reverse transcriptase.

Insert the factor VIII cDNA into a bacterial vector near a promoter site.

Transform the vector into E. coli bacterium.

E.coli expresses factor VIII.

Step-by-step explanation:

This is the way to produce a human protein in a bacterial model, since bacteria does not perform the splicing process is better to generate a cDNA copy of the mRNA, since it has the introns already removed. If you use the gene from the genomic DNA, it will have the exons and the introns, and the bacteria is not able to process the RNA to a mature state.