Bio unit 7 pt 2

1. Molecular Biology Techniques

1.1 DNA Extraction

The process of isolating DNA from an organism (DNA/Org) involves several critical steps to ensure the genetic material is pure for analysis:

  1. Lysis: The cell and nuclear membranes are broken open to release the cellular contents.

  2. Digestion: Specific enzymes, such as protease, are utilized to break down proteins.

  3. Separation: The sample undergoes centrifugation to separate cellular debris from the aqueous DNA.

  4. Precipitation: Cold alcohol is added to the solution to make the DNA visible as it precipitates.

  5. Purification: The resulting DNA strand is washed and resuspended in a buffer.

1.2 polymerase chain reaction (PCR)

The Polymerase Chain Reaction is a vital method used to amplify DNA samples, creating many copies of a specific Gene. This process takes place in a Thermal Cycler.

  • Key Components:

    • DNA Polymerase: An enzyme that synthesizes DNA molecules from deoxyribonucleotides.

    • Primer: A short nucleic acid sequence providing a starting point for DNA synthesis.

  • The Cycle:

    1. Denaturation: Application of heat separates the double-stranded DNA.

    2. Annealing: The temperature is adjusted to allow Primers to bind to the target sequences.

    3. Extension: DNA Polymerase adds nucleotides to the sequence, extending the new strand.

    4. Repetition: The steps are repeated in a Cycle for exponential growth of the sample DNA.

1.3 Gel Electrophoresis

A method used to separate and analyze macromolecules (DNA and proteins) based on their size and electrical charge.

  • Requirements: Agarose Gel, electricity (+, --), and a sample mixed with dye.

  • Process:

    1. Prep: The Agarose Gel is placed into the electrophoresis chamber.

    2. Load: The DNA sample is placed into the ladders or wells at the negative end of the gel.

    3. Apply: An electrical current is applied across the gel.

    4. Monitor: Fragments move through the matrix; smaller pieces move faster than larger ones.

    5. Observation: The gel is viewed under UV light after staining to identify the positions and sizes of the genetic bands.

2. Genetic Engineering and Modification

2.1 The Genetic Modification Process

Genetic Mod is the process of altering the Genome of an organism to create a Transgenic organism.

  1. Isolate: Identify and isolate the specific Gene of Interest.

  2. Cut: A Restriction Enzyme is used to cut the Gene of Interest and the target Plasmid (a small double-stranded DNA molecule).

  3. Ligation: The enzyme Ligase is used to combine the target gene with the Plasmid.

  4. Transformation: The host cell is transformed by inserting the modified genetic material.

  5. Clone/Screen: Cells are grown and screened to identify successful mutations.

2.2 Gene Regulation and Biosynthesis

  • Promoter: A region of DNA that initiates the transcription of a particular gene.

  • Repressor: A protein that inhibits gene transcription by binding to the DNA.

  • Transcription Factor: Proteins that control the rate of transcription of genetic information from DNA \rightarrow RNA.

  • Biosynthetic Path: A series of chemical reactions in a cell that produce complex molecules. This includes the production of pigments such as:

    • Carotenoids: Pigments that are red, yellow, or orange.

    • Anthocyanins: Water-soluble vacuolar pigments that appear red, purple, or blue depending on the pH.

3. Genetics and Breeding Strategies

  • Hybridization: A form of breeding that involves crossing individuals with distinct genetic traits.

  • Inbreeding: The breeding of individuals with identical or highly similar DNA due to a shared ancestor.

  • Bacterial Mutation: A permanent or random change in the nucleotide sequence of a bacterial genome.

  • Polyploid Plants: Plants that contain more than two sets (+2$$) of chromosomes, often leading to increased size or vigor.