Recombinant DNA tech, RNA tech, Protein Expression, Protein tech Assignment

Recombinant DNA tech, RNA tech, Protein Expression, Protein tech - Assignment Example Question Two For a plasmid to act as an effective cloning vector that can transfer the gene of interest into the target cells, it must exhibit the ability to replicate upon entry into the target cells. Moreover, it must have several cloning sites that allow the insertion of the gene of interest. The plasmid must have a defined origin of replication and effective promoters that ensure successful expression of the foreign gene. Question Three Polymerase chain reaction is one of the ways in which the 1kb gene may be quantified. This process yields many copies of the gene of interest, making other analytical processes possible. The modern Polymerase Chain Reaction (PCR) exhibits a high level of automation, and yields multiple copies of the gene. The second way of yielding great amounts would involve insertion of the gene into bacteria. After replication of the plasmid in the bacteria, the gene of interest multiplies. Question Four In order to identify the genes in the human liver whose e xpression occurs only when under pressure, a cDNA library would be the most effective. This library would focus on the fragments that undergo transcription and expression. Construction of such a library would require the sequencing of the genes of interest and inserting them into a plasmid vector. The library would also have reverse transcribed messenger RNAs for the genes, and this would involve the use of reverse transcriptase to yield DNA complementary to the messenger RNA. Question Five Type II Restriction Endonucleases do not degrade bacterial chromosomal DNA because they exhibit specificity for foreign DNA. It would be irrational for these endonucleases to cleave the host DNA. Therefore, they limit their activity to breaking down foreign DNA into fragments but preserving host DNA. Such specificity has enabled geneticists to develop DNA fragmentation techniques using restriction enzymes found in bacteria. Question 6 A double stranded circular DNA with four recognition sites for the HindIII, would be fragmented into four fragments after digestion. The restriction would cleave the circular DNA at the four recognition sites yielding four independent fragments, contrary to the five that would result after linearization of the DNA. The difference would emerge because of the evident practical observation made by geneticists who have highlighted that circular DNA yields one less fragment after digestion with restriction enzyme, compared to linear DNA with the same restriction sites. Question 7 In order to screen a cDNA library, the high-density screening method would prove to be highly effective. This method requires the use of high concentrations plating. The choice of this screening factor would be motivated by the fact that it presents a platform for the geneticist to analyze the different fragments through the visualization on a single plate. The technique also proves effective when the gene under study codes for a specific protein. Part 2 Question 1 In orde r to separate cells obtained from tissues of normal mice compared to those obtained from mice engineered with highly active muscle cells, a specific method of analysis is required. The first step would involve isolation of cells from both tissues. After isolation of those cells, effective culture would follow to prepare the cells for effective analysis. It would be necessary to isolate the mitochondrial DNA and nuclear DNA from both types of cells. Muscle cells have