Aspirant Academy

Science and Technology

Biotechnology and Genetic Engineering

Biology: Cell, Plant Parts, Nutrition/Reproduction, Human Physiology, Food/Nutrition, Immunity/Diseases, Microbes, Fermentation, Biotech/Genetic Engineering, GMO Ethics, Vaccines/CRISPR/mRNA, Artificial Organs

Paper II · Unit 2 Section 8 of 13 0 PYQs 31 min
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Biotechnology and Genetic Engineering

7.1 Recombinant DNA Technology

Steps in Creating Recombinant DNA (rDNA)

  1. Isolate target gene from donor organism (using restriction enzymes to cut at specific sequences).
  2. Cut vector (usually a plasmid or viral DNA) with same restriction enzyme.
  3. Ligate (join) target gene into vector using DNA ligase → recombinant DNA.
  4. Transform recombinant vector into host cell (bacteria, yeast) via heat shock, electroporation.
  5. Clone host cells → all daughter cells contain target gene → express protein of interest.

Key Tools

  • Restriction endonucleases ("molecular scissors"): Cut DNA at specific palindromic sequences. E.g., EcoRI cuts at GAATTC.
  • DNA ligase ("molecular glue"): Joins DNA fragments.
  • PCR (Polymerase Chain Reaction): Amplifies specific DNA sequences millions-fold in vitro using thermostable Taq polymerase. Used in forensics, disease diagnosis (RT-PCR for COVID-19), paternity testing.
  • Gel electrophoresis: Separates DNA fragments by size for analysis.

7.2 Applications of Genetic Engineering

Medicine

  • Recombinant human insulin (Humulin, 1982): Human insulin gene inserted into E. coli → mass production. Replaced animal insulin (pig/cow); no allergic reactions.
  • Human Growth Hormone (somatotropin): Produced in E. coli; treats dwarfism.
  • Tissue Plasminogen Activator (tPA): Clot-busting drug for stroke; recombinant protein.
  • Erythropoietin (EPO): Recombinant protein promoting RBC production; used in anaemia and (controversially) blood doping in sports.

Agriculture — GM Crops

  • Bt crops (Bacillus thuringiensis): bt gene (encoding insecticidal Cry protein) inserted into crop plants. Bt cotton — India's only approved GM food/fibre crop (2002); ~12.3 million hectares (90%+ of India's cotton area). Reduces bollworm infestation.
  • Herbicide-tolerant crops (HT): e.g., Roundup Ready soybean (Monsanto) — tolerates glyphosate herbicide, killing weeds not crop.
  • Golden Rice: Contains β-carotene genes from daffodil and maize; addresses Vitamin A deficiency. Under regulatory review in Philippines, Bangladesh; stalled in India.

India's GMO Regulatory Framework

  • GEAC (Genetic Engineering Appraisal Committee) under MoEF&CC — apex body for approval of GMO research and commercial release.
  • DBT (Department of Biotechnology) — funds biotech research; manages bio-containment laboratory BSL-4 in Pune.
  • Bt brinjal approved by GEAC (2010) but moratorium imposed by Environment Minister Jairam Ramesh; remains under debate.

7.3 CRISPR-Cas9 (PYQ-potential — new in 2026 syllabus)

What is CRISPR?

CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) was discovered as a bacterial immune system that stores fragments of viral DNA to recognise and destroy future viral invasions.

CRISPR-Cas9 as a Gene Editing Tool

Developed by Jennifer Doudna (UC Berkeley) and Emmanuelle Charpentier (Max Planck Institute) — awarded Nobel Prize in Chemistry in 2020.

Mechanism: A guide RNA (gRNA) is designed to match a target DNA sequence. It directs the Cas9 enzyme (a "molecular scissors") to bind and cut double-stranded DNA at the precise target location. The cell's DNA repair mechanisms then either: (a) join the cut ends (disabling the gene) or (b) incorporate a new DNA sequence (gene replacement).

Advantages over Older Gene Editing Tools (TALEN, zinc-finger nucleases)

  • Much simpler and cheaper — gRNA synthesis is straightforward
  • More precise — single-nucleotide precision
  • Multiplexing — can edit multiple genes simultaneously
  • Faster — weeks instead of months/years

CRISPR Applications

  • Disease treatment: Sickle cell disease — CASGEVY (CTX001, CRISPR Therapeutics) became the first CRISPR-based therapy approved by FDA (December 2023) for sickle cell disease and beta-thalassaemia.
  • Cancer therapy: Engineering T cells (CAR-T cells) with CRISPR to better recognise and kill cancer.
  • Agriculture: Disease-resistant crops (CRISPR-edited mushrooms that don't brown; high-yield wheat).
  • COVID-19 diagnostics: SHERLOCK (CRISPR-based) detection system — CSIR-IGIB's FelUDA (FNCAS9 Editor Linked Uniform Detection Assay) — India's CRISPR-based COVID test approved by DCGI in 2021.

Ethical Concerns

  • Germline editing: CRISPR changes in embryos are heritable — He Jiankui (China, 2018) controversially created the world's first genome-edited babies (CCR5 gene modified to confer HIV resistance) — condemned globally as unethical; sentenced to 3 years imprisonment.
  • Designer babies and the enhancement vs therapy distinction.
  • Environmental risks of gene drive (engineered self-spreading genetic changes in wild populations — e.g., to eliminate malaria mosquitoes).
  • Inequitable access if only wealthy nations can afford gene therapies.