Define transcription. Explain the process of transcription in bacteria with suitable diagram.

The genetic information flows from DNA to mRNA and then to the protein-synthesizing machinery. The process of synthesis of RNA strands from the DNA is known as transcription. In a prokaryotic cell, this takes place in the cytoplasm as they lack a membrane-bound nucleus. It is a process coupled with the translation in a prokaryotic cell. A transcription unit in prokaryotic cells is the specific region of DNA that undergoes this process catalyzed by DNA-dependent RNA polymerase enzyme. The complete process of transcription requires: Ribonucleotides (NTPs)- Adenine, Uracil (in place of Thymine), Guanine, Cytosine RNA polymerase enzyme (it has different subunits) and other transcription factors. Template DNA strand- Only a specific region (gene) is transcribed in a 3' to 5' direction and not the entire genome. Another strand of the dsDNA will be complementary to the newly synthesized mRNA transcript and would not undergo the transcription process. Transcription in a prokaryotic cell involves three steps: Initiation– The dsDNA is unwounded to single-stranded forms near the initiation site to provide a template strand. The DNA-dependent RNA polymerase (RNAP) binds to the specific sequence on the template strand named promoter sequence (Pribnow box / TATA box) and initiates the transcription of the template strand. The sigma factors of RNAP enzyme help them to bind the promoter sequence. Elongation– After the synthesis of about 10 base pairs of long RNA, the sigma factor of RNAP enzyme slides over from the promoter site over the template DNA strand in a 5’ – 3’ direction continuously all the while polymerizing NTPs to form mRNA strand. Termination– This is the last stage that terminates the transcription process. There are two mechanisms: Rho-independent transcription termination– transcription stops due to a specific sequence in terminator DNA. The terminator DNA contains intrachain self-complementary sequences in the newly formed transcript RNA that form the hairpin structure that hinders the RNAP enzyme and pauses its function. This may result in the unloading of the RNAP enzyme from the DNA template. Rho-dependent transcription termination – transcription is terminated by rho protein that recognizes the DNA sequences act as a signal of termination. It binds to RNA after the translation has completed, but transcription is still going on. This factor causes the dissociation of the RNAP enzyme from the DNA template.

An agent, such as a chemical or radiation, that could induce mutation are called mutagen. Mutations are sudden heritable changes that could be natural or artificial induced. An induced mutation is of great importance in crop improvement.

Explain briefly the process of translation in prokaryotes.

The process of translation in prokaryotes: Translation refers to the process of polymerization of amino acids to form a polypeptide or a protein. In prokaryotes, translation takes place in the cytoplasm. The order and sequence of amino acids are defined by the sequence of bases in the mRNA and the amino acids are joined by a bond which is known as a peptide bond. The formation of a peptide bond requires energy and thus, amino acids are activated in the presence of ATP and linked to their cognate tRNA by the process of charging of tRNA or aminoacylation of tRNA. If charged tRNA are brought close enough, a peptide bond is formed that is enhanced by the presence of a catalyst such as a ribosome. The ribosome in its inactive state exists as two subunits; a large subunit (50S) and a small subunit (30S). There are two sites in the large subunit, for subsequent amino acids to bind to and thus, become close enough to each other for the formation of a peptide bond. A translational unit in mRNA is the sequence of RNA that is flanked by the start codon (AUG) and the stop codon and codes for a polypeptide. An mRNA also has some additional sequences that are not translated and are referred to as untranslated regions (UTR). The UTRs are present at both 5'-end (before start codon) and at 3'-end (after stop codon) which are required for an efficient translation process. After activation of amino acids, translation starts with its three steps: Initiation Elongation Termination Initiation: For initiation, the 70S ribosome binds to the mRNA at the start codon (AUG) which is recognised only by the initiator tRNA. Elongation: The ribosome proceeds to the elongation phase of protein synthesis. During the elongation stage, complexes composed of an amino acid linked to tRNA, sequentially bind to the appropriate codon in mRNA by forming complementary base pairs with the tRNA anticodon. The ribosome moves from codon to codon along the mRNA. Amino acids are added one by one, translated into polypeptide sequences dictated by DNA and represented by mRNA. Termination: In the end, a release factor binds to the stop codon, terminating translation and releasing the complete polypeptide from the ribosome.

Describe the mechanism of DNA replication with a suitable diagram.

The process of DNA replication takes place as follows: It begins at a point called the origin of replication (ORI), to form a replication fork. The separated strands act as templates for the synthesis of new strands. DNA replicates in the 5' -→ 3' direction. dNTPs (deoxyribonucleoside-triose phosphates) act as substrates and also provide energy for the polymerization of nucleotides. DNA polymerase is an enzyme that assembles a new DNA strand that is complementary to the template strand. DNA polymerase continues to move along the template strand and add new nucleotides to the growing or complement any strand until the entire genome is replicated. The DNA polymerase forms one new strand (leading strand) in a continuous stretch in the 5' -→ 3' direction (continuous synthesis). The other new strand is formed in small stretches (Okazaki fragments) in the 5' -→ 3' direction (discontinuous synthesis). The Okazaki fragments are then joined together to form a new strand by an enzyme, DNA ligase. This new strand is called the lagging strand.