The Molecular Word Processor: The Next Step Beyond CRISPR

CRISPR is a powerful gene-editing tool that works like molecular scissors. It can cut DNA and help scientists edit genes. But CRISPR has one problem: it makes a full cut in the DNA, and sometimes the cell repairs this cut in the wrong way. This can lead to mistakes.

Because of this, scientists have created newer and safer gene-editing tools. These new tools work like a “Molecular Word Processor” instead of scissors. They can edit DNA without making big cuts. This makes gene editing more accurate and less risky.

CRISPR-Cas9: The Molecular Scissors

CRISPR-Cas9 was the first major gene-editing tool.

How it works:

• A guide RNA finds the place in the DNA that needs editing.
• The Cas9 enzyme cuts both strands of the DNA.
• The cell repairs the cut, but sometimes the repair is messy or wrong.

This method is powerful but not always safe because it creates a double-strand break.

Base Editing: The Molecular Pencil

Base Editing was the next improvement. It does not cut the whole DNA. Instead, it changes just one letter of the DNA.

How it works:

• A modified Cas9 makes a small nick in one DNA strand.
• An enzyme changes one DNA base to another (like C to T or A to G).

Why it matters:
Many diseases are caused by a single wrong DNA letter. Base editing can fix these “DNA typos” safely.

Prime Editing: The Molecular Word Processor

Prime Editing is the most advanced tool so far. It works like the “Find and Replace” feature in a computer.

It can:

• Change any DNA letter
• Insert small DNA sequences
• Delete small DNA sequences
   all without making a dangerous double-strand cut.

Prime Editing uses three parts:

1. Nickase Cas9 – cuts only one DNA strand
2. Reverse Transcriptase – writes the new DNA
3. pegRNA – guides the editor and provides the corrected DNA sequence

Steps:

1. The pegRNA guides the editor to the target.
2. Nickase Cas9 cuts one strand.
3. The Reverse Transcriptase writes the new DNA.
4. The cell replaces the old DNA with the new, corrected version.

This method is clean, safe, and very accurate.

Why Prime Editing Is Important

Prime Editing can do:

• Substitutions (change one base)
• Insertions (add small pieces)
• Deletions (remove small pieces)

It can fix around 89% of known genetic mutations.
And it does this without making a full cut in the DNA.

Future Tools: Transposons and TIGR

Scientists are working on even newer systems:

1. Transposon-based editors
   These may help insert large pieces of DNA, like whole genes.
2. TIGR systems
   These are very small proteins that may edit any part of the genome easily.

These tools could make gene editing even more powerful and flexible.

Prime Editing and Sickle Cell Disease

Sickle Cell Disease happens because of one small mistake in the HBB gene.

• Healthy DNA: GAG
• Sickle DNA: GTG

This single-letter change causes red blood cells to become sickle-shaped.

Prime Editing can correct GTG back to GAG by changing just one base.

Why it is better than CRISPR-Cas9:

• CRISPR makes a full cut; Prime Editing makes only a small nick
• Prime Editing is more accurate
• Prime Editing causes fewer mistakes

How treatment works:

1. Doctors collect the patient’s blood stem cells.
2. They edit the cells using Prime Editing.
3. The corrected cells are placed back into the patient.
4. These cells can make healthy red blood cells for life.

This could be a permanent cure.

The Future of Gene Editing

CRISPR started the revolution.
Base Editing made it cleaner.
Prime Editing made it precise.

Soon, gene editing may cure diseases, help in drug discovery, and prevent genetic problems before they occur.

We are entering a future where DNA can be edited as easily as editing words in a document.

Table: Major Fields of Biotechnology and Their Key Applications

Biotech Field	Key Focus	Important Applications	Examples
Medical Biotechnology	Developing therapies & diagnostics	Vaccine development, gene therapy, disease diagnostics	mRNA vaccines, CRISPR-based therapies
Agricultural Biotechnology	Crop improvement & protection	High-yield crops, pest-resistant plants, stress-tolerant varieties	Bt Cotton, Golden Rice
Industrial Biotechnology	Large-scale bio-processing	Enzyme production, biofuels, bioplastics	Bioethanol, biodegradable plastics
Environmental Biotechnology	Pollution control using microbes	Wastewater treatment, bioremediation, bio-filtering	Oil spill clean-up, heavy metal removal
Food Biotechnology	Food quality & preservation	Fermentation, probiotic foods, nutritional enhancement	Yogurt, cheese, fortified foods
Marine Biotechnology	Exploring marine organisms	Novel enzymes, pharmaceuticals, bioactive compounds	Anticancer compounds from algae
Microbial Biotechnology	Using microbes for innovation	Antibiotics, enzyme production, fermentation industries	Penicillin, lactase enzyme

 

Advanced Biotechnology Quiz

15 MCQs for CSIR-NET / GATE / DBT-JRF practice. Choose one answer for each question, then click Submit.

Q1. In Sanger sequencing, which molecule acts as a chain terminator causing termination of DNA synthesis?

A. dATP B. ddNTP (dideoxynucleotide) C. rNTP D. DNA ligase

Q2. Which of the following is FALSE about quantitative PCR (qPCR) using SYBR Green dye?

A. SYBR Green binds to double-stranded DNA. B. SYBR Green can differentiate between specific and non-specific products. C. Melt curve analysis helps identify primer-dimers. D. SYBR Green fluorescence increases proportionally with product amount.

Q3. In enzyme kinetics, which plot is used to determine Vmax and Km by linearization?

A. Michaelis-Menten plot (v vs [S]) B. Lineweaver-Burk plot (1/v vs 1/[S]) C. Scatchard plot D. Hill plot

Q4. Which method is most suitable for isolating high-molecular-weight genomic DNA free from RNA contamination?

A. Boiling lysis without RNase B. CTAB extraction followed by RNase treatment C. Phenol-only extraction D. Direct PCR from colony

Q5. In plant tissue culture, which plant growth regulator combination is commonly used to induce callus formation?

A. High cytokinin, low auxin B. High auxin, low cytokinin C. No hormones D. Gibberellin alone

Q6. Which technique is used to measure genome-wide DNA methylation at single-base resolution?

A. ChIP-seq B. ATAC-seq C. Bisulfite sequencing D. RNA-seq

Q7. In next-generation sequencing, 'paired-end' reads are useful because they:

A. Reduce sequencing error rate to zero B. Provide sequence information from both ends of a DNA fragment, helping with alignment across repeats C. Double the read length automatically D. Only used in Sanger sequencing

Q8. Which statement about Agrobacterium tumefaciens-mediated transformation is CORRECT?

A. It is effective only in monocots and not dicots B. It transfers T-DNA from the Ti plasmid into the plant genome C. It uses electroporation to enter plant cells D. It inserts RNA directly into the nucleus

Q9. Which of the following best describes a 'silent mutation' at the codon level?

A. Changes an amino acid to a stop codon B. Changes the DNA sequence but not the encoded amino acid C. Always causes a frameshift D. Deletes an exon

Q10. In fermentation, which parameter is most directly used to estimate microbial biomass in real time?

A. Optical density (OD) at 600 nm B. pH measurement C. Glucose concentration D. CO2 sensor only

Q11. Which DNA repair pathway is primarily active during non-dividing (G0/G1) cells and is error-prone?

A. Homologous recombination (HR) B. Base excision repair (BER) C. Non-homologous end joining (NHEJ) D. Mismatch repair (MMR)

Q12. Which viral vector is most commonly used for stable integration into dividing mammalian cells for gene therapy?

A. Adeno-associated virus (AAV) B. Adenovirus C. Retrovirus / Lentivirus D. Bacteriophage

Q13. Which technique separates proteins based on isoelectric point and molecular weight sequentially?

A. SDS-PAGE B. 2D gel electrophoresis C. Western blot D. Size-exclusion chromatography

Q14. In population genetics, which effect describes loss of genetic variation when a new population is founded by a small number of individuals?

A. Bottleneck effect B. Founder effect C. Gene flow D. Stabilizing selection

Q15. Which reagent is commonly used as a reducing agent to break disulfide bonds during SDS-PAGE sample preparation?

A. SDS B. β-mercaptoethanol (or DTT) C. EDTA D. Triton X-100

Tip: This quiz is for practice. For exam-style preparation, attempt without notes and time yourself for 20 minutes.