To synthesize rRNA in a test tube using DNA isolated from mouse cells, in addition to the template DNA, ribonucleotides, and the necessary transcription factors, you should add RNA polymerase I.
To synthesize rRNA (ribosomal RNA) in a test tube using DNA as a template, you would need to add RNA polymerase I to the reaction mixture. RNA polymerase I is responsible for transcribing rRNA genes into precursor rRNA, which is then processed into the mature rRNA used to form the ribosomal subunits involved in protein synthesis. Thus, the correct answer is c. RNA polymerase I.
To synthesize ribosomal RNA (rRNA) in a test tube using DNA isolated from mouse cells, indeed RNA polymerase I would need to be added alongside the template DNA, the ribonucleotide triphosphates (which are the building blocks of RNA), and the necessary transcription factors. RNA polymerase I is responsible for the transcription of rRNA in the cell nucleus of eukaryotes, excluding the 5S rRNA, which is transcribed by RNA polymerase III.
Here are the logical steps you would follow for the in vitro (test tube) synthesis of rRNA:
1. Isolation of Template DNA - You would begin by isolating the DNA from mouse cells, ensuring that the regions coding for rRNA are present.
2. Preparation of Transcription Mix - Prepare a reaction mix containing: - The isolated DNA as a template. - A mixture of all four types of ribonucleotide triphosphates (ATP, UTP, GTP, CTP). - The necessary transcription factors specific to RNA polymerase I. These factors help the polymerase to recognize the start site of rRNA genes and regulate the process of transcription. - Magnesium ions and a buffer to maintain an appropriate pH and salt concentration for enzyme activity.
3. Adding RNA Polymerase I - Introduce RNA polymerase I into the reaction mix. In eukaryotes, there are three main types of RNA polymerases (I, II, III), each responsible for transcribing different kinds of genes. RNA polymerase I is specialized in transcribing rRNA genes.
4. Incubation - Allow the reaction to proceed by incubating the mix at an appropriate temperature for the enzyme to function efficiently, typically around 37°C (the body temperature of mice).
5. Termination - Once sufficient time has passed for transcription to occur, stop the reaction. This can be done by using a method that inactivates the RNA polymerase, such as the addition of a chelating agent that removes the magnesium ions or heating the mixture to denature the enzyme.
6. Analysis - Analyze the synthesized RNA using appropriate methods such as gel electrophoresis or Northern blotting to confirm the synthesis and integrity of the rRNA.