3.5 Genetic Modification and Biotechnology

Polymerase Chain Reaction (PCR)

**Artificial** method of replication used to rapidly **amplify DNA sequences.** Involves three key steps (which are repeated):

* **Denaturation** – Heat separates strands (no helicase).
* **Annealing** – Primers designate copying sequence.
* **Elongation** – Taq polymerase copies the sequence

Gel Electrophoresis

Separates **proteins** or **fragments** of DNA according to their size.

* DNA fragments are placed in an agarose gel.
* Proteins are placed in a polyacrylamide gel.

DNA Profiling

Comparison of DNA

DNA Profiling: Forensics

* DNA sample collected.
* STR loci selected.
* **PCR amplification.**
* **Gel electrophoresis.**

\
Suspect and sample should be a complete match

DNA Profiling: Paternity

Children inherit DNA from **both parents**, all of the **child’s bands** should match to either the mother or father (i.e. **combination of parents**)

Gene Transfer

Process by which genes from one organism are inserted into another organism (i.e. horizontal gene transfer).

* Genetic code is *universal.*

What is a **vector**?

Molecular **vehicle** used to artificially **carry genetic material into a host cell.**

* Plasmids: **Circular DNA** molecules capable of **autonomous gene expression.**
* Viruses: **Inject** their **genetic material** directly into **host cells.**

Gene transfer key steps

**Four** key steps:

* **DNA extraction** – **Gene** of interest **and vector** (*virus or plasmid*) are ==**isolated.**==
* **Digestion** – Gene and vector are **cut** with restriction endonucleases.
* **Ligation** – **Gene** of interest is **placed into the vector.**
* **Transformation** – Recombinant vector **inserted into host cells.**

Step 1: DNA Extraction

* Gene of interest **isolated** from organism.
* Gene is amplified using **PCR** (along with a plasmid).

Step 2: Digestion and Ligation

* **Plasmid** and **gene** **cut** with a specific restriction enzyme.
* Gene is spliced into plasmid vector by **DNA ligase**.

Step 3: Transformation and Expression

* **Recombinant plasmid** is **inserted into a host cell.**
* Antibiotic selection may be used to select for successful transgenic cells (if plasmid has an antibiotic resistance gene).
* **Transgenic cells express new protein** (for extraction / use)

Clones

Groups of **genetically identical** organisms (derived from **single parent cells**).

* **Naturally** via ==__**a**__sexual reproduction.==
* **Artificially** via ==embryonic division== or **somatic** cell nuclear transfer.

Natural Cloning: Animals

Certain species of animals can clone **themselves**.


1. Binary fission: **divides** **equally** in two.
2. Budding: cells **split off** from parent organism.
3. Fragmentation: new organism **grows** from fragment.

Natural Cloning: Plants

Plants have the capacity for **vegetative propagation**, small parts of the plant **grow independently.**

* **Stem cuttings** are separated portions of plant stems that can **regrow into new, independent clones.**

Natural Cloning: Humans

Identical twins (**monozygotic**) are created when a **fertilised egg splits into two** separate cell groups

Artificial Cloning

Using differentiated adult tissue called **somatic cell nuclear transfer (**@@**SCNT**@@**)**

* **Reproductive** cloning – Produces **new** __**organisms**__ (%%repopulate%% endangered species)
* **Therapeutic** cloning – Produces **new** __**tissue**__ (%%transplantations / disease treatments%%)

Artificial Cloning: SCNT

Cloning **embryos** from **differentiated adult tissues:**

* Nucleus is removed from adult body cell.
* Egg cell nucleus destroyed by UV radiation.
* Body cell nucleus fused to enucleated egg.
* Electrical current stimulates egg to divide.
* Developing embryo implanted in surrogate.
* Embryo develops into a cloned animal.