BCHM First Year Revision

Introduction

  • Sinead introduces the lecture concerning molecular and cellular biology and genetics.

  • Basics covered include familiarity with the content and goals of today’s lecture.

  • The lecture is split into two parts: summarizing molecular and cellular biology (Part 1) and discussing the practical aspect of the module (Part 2).

Molecular and Cellular Biology

Central Dogma of Biology

  • Definition: The central dogma describes the directional flow of genetic information.

    • DNA is transcribed into messenger RNA (mRNA).

    • mRNA is translated into protein.

  • Key Concepts:

    • Genes are located on DNA.

    • Genes encode information detailing how to make specific proteins.

Role of mRNA

  • mRNA acts as a bridge between DNA information and protein synthesis.

    • Transcription: Process of synthesizing mRNA from DNA.

    • Translation: Process of synthesizing proteins using the information in mRNA.

Structure of DNA

  • Double-Stranded Helix: DNA is double stranded in a twisted ladder-like formation.

    • Nucleotides: Basic units making up DNA, consisting of:

    • Sugar

    • Phosphate

    • Base group (A, T, C, G)

    • Sugar Phosphate Backbone: Formed by alternating sugar and phosphate groups.

Base Pairing in DNA

  • Two strands of DNA are linked through complementary base pairing:

    • Adenine (A) pairs with Thymine (T) using double hydrogen bonds.

    • Cytosine (C) pairs with Guanine (G) using triple hydrogen bonds.

  • DNA strands run anti-parallel:

    • 5' end: terminal phosphate group

    • 3' end: terminal hydroxyl group

Transcription and mRNA

Role of Template and Coding Strands

  • Coding Strand: The top strand (5' to 3'), also called sense or non-template strand.

  • Template Strand: The bottom strand (3' to 5'), used to transcribe mRNA.

  • Enzyme Role: RNA polymerase binds to the promoter and synthesizes mRNA from the template strand, adding ribonucleotides complementary to the template strand.

Characteristics of mRNA

  • mRNA is complementary to the DNA’s template strand.

  • Differences in base pairs:

    • Thymine in DNA is replaced by Uracil in RNA.

  • Ends are labeled 5' and 3' for identification.

Translation into Protein

Understanding Codons

  • Codon: Sequence of three nucleotides on mRNA that specifies an amino acid.

  • Importance of codons in protein synthesis:

    • The information in codons is translated to form proteins.

  • Start Codon: AUG, codes for Methionine and signifies the start of translation.

  • Stop Codons: Signify termination of translation (Example codons: UAA, UAG, UGA).

Role of Transfer RNA (tRNA)

  • Function: tRNA transports specific amino acids to ribosomes during translation.

  • Anticodon: Region in tRNA that pairs with the complementary mRNA codon, ensuring the correct amino acid is added to the growing peptide chain.

Practical Application of Concepts

Example from Past Paper (2015)

  • Transcription of DNA to mRNA: Identify the coding and template strands to transcribe correctly.

  • Implications of finding start codon and stop codon in transcribed mRNA.

    • Importance of understanding locations of start and stop codons in coding.

Practical Session on Recombinant DNA Technology

Overview

  • The process involves altering DNA from different organisms, using genetic engineering to create recombinant DNA molecules.

  • Genes from various organisms can be combined, leading to new proteins and traits.

Steps of Recombinant DNA Technology

  1. Identify gene of interest.

  2. Cut the gene out and insert it into plasmids using restriction enzymes.

  3. Bacterial Cells: Introduce plasmids to bacteria which replicate quickly.

  4. Isolation of Recombinant Plasmids: Ensures desired proteins can be expressed and harvested.

Plasmids and Restriction Enzymes

  • Plasmids: Circular DNA molecules used in genetic engineering.

    • Plasmids replicate independently from bacterial chromosomes and carry a few genes reducing interference.

  • Restriction Enzymes: Molecular tools that cut DNA at specific sequences.

    • EcoRI example is discussed; it cuts at the GAA TTC sequence, creating sticky ends for joining DNA pieces together.

DNA Ligase Role

  • DNA Ligase: Enzyme that joins DNA fragments by sealing gaps left after sticky end pairing.

Gel Electrophoresis

  • Process: Used to separate DNA fragments by size.

    • A loading gel with wells where DNA samples are inserted and an electric current is applied.

    • Small fragments move faster than larger ones due to pore constraints of the gel.

  • Detection: Use of fluorescent dyes under UV light to visualize separated DNA.

Practical Example: Human Growth Hormone Production

  • Modification Steps:

    • Finding and isolating target gene.

    • Cutting DNA and preparing plasmids.

    • Transformation of bacteria to produce desired protein (growth hormone).

Transformation Process

  1. Bacterial cells are made competent to uptake plasmids.

  2. Incubation of bacterial cells with plasmids for recovery and replication.

Lab Activities

  • The practical entails cloning a gene for red fluorescent protein into a plasmid and inserting into bacterial cells.

  • Success measured by the color change in bacteria expressing the new protein.

Challenge Question

  • Acknowledgment of an extra challenge question requiring advanced chemistry knowledge, deemed optional for students.

  • Support offered for students needing additional clarification.

Conclusion

  • Sinead invites questions and expresses readiness to assist during practical sessions.