New Biotechnology

Discussion on different methods of altering plant and organism characteristics
  - Conventional breeding involves selective breeding techniques
  - Genetic modification (GM) involves altering an organism's DNA by transferring genes from one organism to another.
    - Example crops include virus-resistant plants and high-yield crops.
  - Genome editing allows precise modifications directly to an organism’s DNA.

Genetic Modification:
- A technique to change an organism's traits by transferring DNA from one organism to a different organism.
- Commonly employs methods like Agrobacterium tumefaciens for gene insertion.
Genome Editing:
- Refers to a suite of technologies that allow the direct modification of an organism's DNA.

When using Agrobacterium tumefaciens, the process follows:
  1. Insertion of a gene for the desired trait into the Ti plasmid.
  2. Creation of recombinant Ti plasmid that can integrate into plant cells in culture.
  3. Regeneration of a plant from the modified plant cells, which contains the new gene integrated into its chromosome.

Recombinant DNA:
- Requires a mechanism for stable integration into the host's genome.
- Transposons (transposable elements) are DNA pieces that can move within the genome.

Sense vs. Antisense DNA:
  - Sense (Coding strand): The strand of DNA that is transcribed into mRNA.
  - Antisense (Non-coding strand): Template strand for transcription.
  - Example:
    - Sense strand: 5' ATGTGGCAAGTGCCCCC 3'
    - Antisense strand: 3' TACACCGTTCACGGGGTA 5'.

Knockin (KI):
- Insertion of a gene at a specific locus in the genome, often using CRISPR-Cas9 or homologous recombination.
- Goals include replacing a gene or tagging it for study.
Knockout (KO):
- A technique that completely inactivates a gene, resulting in a loss of function.
Knockdown (KD):
- Reduces expression of a gene, resulting in a temporary phenotypic alteration.
Overexpression (OE):
- Increases gene expression abnormally, typically achieved through strong promoter use or multiple gene copies.

CRISPR-Cas9:
  - A genome editing tool that allows for precise cuts to DNA and repair processes.
  - Components include:
    - Guide RNA (gRNA): Binds to Cas9 and directs the cut location in DNA.
    - Cas9 Enzyme: Cuts the DNA at the specified location given by gRNA.
Homologous Recombination (HR) and Nonhomologous End Joining (NHEJ):
- Two repair methods after DNA has been cut by Cas9. HR can insert new DNA, while NHEJ can create small mutations.

RNA interference is a biological process in which RNA molecules suppress gene expression.
Mechanism:
1. dsRNA cuts into smaller fragments by Dicer enzyme.
2. Fragments guide RISC complexes to target mRNA for degradation.
Applications include:
- Gene knockdown and functional genomics.
- Used in medicine, biotechnology, food enhancement, and transgenic plants.

Papaya ringspot virus (PRSV) Resistance:
- Genetic modification of papaya using RNAi to confer resistance against PRSV (Jia et al. 2017).
- Study highlighting RNAi's role in resistance to viral infections.

Sickle Cell Anemia and β-Thalassemia Treatment:
- The use of CRISPR-Cas9 gene editing to modify genetic expressions related to these disorders (Frangoul et al. 2021).
- Highlights the practical applications of gene editing in treating genetic conditions.

Stages in using CRISPR-Cas9:
  1. Acquisition of foreign DNA: CRISPR memories of previous infections are maintained.
  2. RNA Processing: Input of RNA guides to target DNA.
  3. Targeting and Cutting: Cas9 enzyme cuts the DNA at specific sites guided by RNA sequences.

Continuous exploration of RNAi and CRISPR technologies in gene editing and functional genomics.
Development of new crops and treatments for genetic disorders through advanced gene editing techniques.