CRISPR-Cas Systems — MCQs
Which is the correct sequence of steps in isolating desirable protein using recombinant DNA technology? 1. Expression of protein and lysis of the bacterial cell 2. Incorporation of genes into bacteria 3. SDS PAGE 4. Protein elution 5. Column chromatography
Which of the following statements best describes the mechanism of action of insulin on target cells?
Which of the following is the platinum-based chemotherapeutic agent used as first-line treatment for ovarian carcinoma?
Which of the following statements about gene therapy is false?
Which of the following is used to detect abnormal gene sequences EXCEPT?
Which type of mutation can act as a suppressor to restore the wild-type phenotype in organisms carrying a mutant gene?
Which malformation is associated with mutations in the HOX gene?
A 15-year-old boy returned from a vacation to his relatives' village. He developed a severe headache and purulent nasal discharge and was diagnosed with primary amebic meningoencephalitis (PAM). He died 5 days later. Which is the most likely etiological agent?
Transmission of R factor is by which mechanism?
Resistance to drugs in tuberculosis develops by which mechanism?
CRISPR-Cas Systems Indian Medical PG Practice Questions and MCQs
Question 1: Which is the correct sequence of steps in isolating desirable protein using recombinant DNA technology? 1. Expression of protein and lysis of the bacterial cell 2. Incorporation of genes into bacteria 3. SDS PAGE 4. Protein elution 5. Column chromatography
- A. 2,1,3,5,4 (Correct Answer)
- B. 2,4,5,3,1
- C. 1,2,4,3,5
- D. 1,5,2,4,3
Explanation: ***2,1,3,5,4*** - This sequence accurately reflects the typical order of operations in **recombinant protein isolation**: first, the gene is introduced into bacteria, then protein is expressed and cells lysed, followed by **SDS-PAGE as an intermediate quality check** to confirm protein expression before proceeding to purification steps (column chromatography and elution). - The process starts with gene incorporation, includes an analytical checkpoint after lysis, and ends with purified protein elution. *2,4,5,3,1* - This sequence is incorrect because **protein elution (4)** and **column chromatography (5)** are purification steps that occur *after* protein expression and cell lysis. - **Lysis (1)** cannot happen after elution, as cells must be lysed first to release the protein for purification. *1,2,4,3,5* - This sequence is incorrect because **expression and lysis (1)** must occur *after* the gene has been **incorporated into bacteria (2)** - the gene must be present before it can be expressed. - Additionally, **protein elution (4)** should follow **column chromatography (5)**, as elution is the step where protein is collected from the chromatography column. *1,5,2,4,3* - This sequence is incorrect because **incorporation of genes (2)** must be the first step - the gene needs to be in the bacteria before any expression, lysis, or purification can occur. - Starting with **expression and lysis (1)** before gene incorporation is impossible.
Question 2: Which of the following statements best describes the mechanism of action of insulin on target cells?
- A. Insulin binds to a receptor on the outer surface of the plasma membrane, activating adenylate cyclase through the Gs protein.
- B. Insulin binds to a cytoplasmic receptor and is transferred as a hormone receptor complex to the nucleus to modulate gene expression.
- C. Insulin enters the cell and causes the release of calcium ions from intracellular stores.
- D. Insulin binds to a transmembrane receptor on the outer surface of the plasma membrane, activating the tyrosine kinase in the cytosolic domain of the receptor. (Correct Answer)
Explanation: ***Insulin binds to a transmembrane receptor on the outer surface of the plasma membrane, activating the tyrosine kinase in the cytosolic domain of the receptor.*** - **Insulin** is a **peptide hormone** and cannot freely pass through the lipid bilayer, thus it binds to a **transmembrane receptor** on the cell surface. - This binding leads to the activation of the receptor's intrinsic **tyrosine kinase activity** in the intracellular domain, initiating a signaling cascade. *Insulin binds to a cytoplasmic receptor and is transferred as a hormone receptor complex to the nucleus to modulate gene expression.* - This mechanism describes the action of **steroid hormones**, which are lipid-soluble and can cross the cell membrane, binding to **intracellular receptors**. - **Insulin** acts via a **cell surface receptor** and its downstream effects are mediated through signal transduction pathways, not direct nuclear translocation. *Insulin binds to a receptor on the outer surface of the plasma membrane, activating adenylate cyclase through the Gs protein.* - This mechanism is characteristic of **G-protein coupled receptors (GPCRs)**, which activate or inhibit enzymes like adenylate cyclase via G-proteins to produce second messengers like cyclic AMP. - The **insulin receptor** is a **receptor tyrosine kinase**, not a GPCR, and does not directly activate adenylate cyclase via Gs protein. *Insulin enters the cell and causes the release of calcium ions from intracellular stores.* - While some hormones and neurotransmitters can trigger the release of intracellular **calcium ions**, this is typically mediated by specific pathways (e.g., GPCRs linked to phospholipase C). - **Insulin** does not directly enter target cells to cause calcium release; its actions are primarily mediated through receptor tyrosine kinase signaling pathways.
Question 3: Which of the following is the platinum-based chemotherapeutic agent used as first-line treatment for ovarian carcinoma?
- A. Cyclophosphamide
- B. Methotrexate
- C. Cisplatin (Correct Answer)
- D. Dacarbazine
Explanation: ***Cisplatin*** - **Cisplatin** is a platinum-based chemotherapy drug that forms **DNA cross-links**, inhibiting DNA synthesis and leading to the death of rapidly dividing cells, making it highly effective against **ovarian carcinoma**. - It is a cornerstone of chemotherapy regimens for ovarian cancer, often used in combination with other agents such as paclitaxel. *Methotrexate* - **Methotrexate** is an **antimetabolite** that inhibits dihydrofolate reductase, thereby interfering with DNA synthesis. - While it is used in various cancers like leukemia, lymphoma, and some solid tumors (e.g., breast cancer, gestational trophoblastic disease), it is **not a primary recommended drug for ovarian carcinoma**. *Cyclophosphamide* - **Cyclophosphamide** is an **alkylating agent** that causes DNA damage, leading to cell death. - It is used in many cancers, including lymphoma, breast cancer, and some leukemias, but it is **not a first-line or primary agent for ovarian carcinoma** in contemporary treatment guidelines. *Dacarbazine* - **Dacarbazine** is an **alkylating agent** primarily used in the treatment of **malignant melanoma** and Hodgkin lymphoma. - It is **not indicated for the treatment of ovarian carcinoma**.
Question 4: Which of the following statements about gene therapy is false?
- A. Gene also considered as drug
- B. Gene therapy can be used to treat some cancers.
- C. Has been tried in cystic fibrosis
- D. Gene therapy is only used for genetic disorders. (Correct Answer)
Explanation: ***Gene therapy is only used for genetic disorders.*** - This statement is **false** because gene therapy has applications beyond just genetic disorders. It is also being explored and used in the treatment of acquired diseases such as **cancer** and **infectious diseases**. - While it's a prominent approach for correcting genetic defects, its scope is much broader, involving the introduction or modification of genes to achieve a therapeutic effect in various conditions. *Gene also considered as drug* - This statement is **true**. Gene therapy products are often regulated as **drugs** or **biological products** by regulatory bodies like the FDA. - This is because they involve the delivery of genetic material that acts to modify gene expression or cell function to produce a therapeutic effect, similar to how traditional drugs work. *Has been tried in cystic fibrosis* - This statement is **true**. Gene therapy has been extensively investigated as a potential treatment for **cystic fibrosis (CF)**. - CF is caused by mutations in the **CFTR gene**, and researchers have attempted to deliver functional copies of this gene to the affected cells, particularly in the lungs, to correct the underlying defect. *Gene therapy can be used to treat some cancers.* - This statement is **true**. Gene therapy is an active area of research and treatment for various **cancers** [1]. - Approaches include introducing genes that make cancer cells more susceptible to chemotherapy, enhancing the immune system's ability to fight cancer, or directly killing cancer cells through gene delivery [1].
Question 5: Which of the following is used to detect abnormal gene sequences EXCEPT?
- A. RFLP analysis
- B. Pyrosequencing
- C. Flow cytometry (Correct Answer)
- D. FISH
Explanation: ***Flow cytometry*** - **Flow cytometry** is primarily used to analyze **cell populations** based on their physical and biochemical characteristics (e.g., size, granularity, and protein expression) by passing them single file through a laser beam, not for direct gene sequencing. - It detects and quantifies cells labeled with **fluorescent antibodies**, making it useful for immunophenotyping, cell sorting, and DNA content analysis, but not for identifying specific gene sequences or mutations. *RFLP analysis* - **Restriction fragment length polymorphism (RFLP) analysis** detects variations in **DNA sequences** by using **restriction enzymes** to cut DNA at specific sites. - Differences in fragment lengths indicate **polymorphisms** or **mutations** within the recognition sites, thereby identifying abnormal gene sequences. *Pyrosequencing* - **Pyrosequencing** is a method of **DNA sequencing** that determines the sequence of nucleotides by detecting the release of pyrophosphate during DNA synthesis. - It is used to identify **single nucleotide polymorphisms (SNPs)** and **short genetic variations**, making it suitable for detecting abnormal gene sequences. *FISH* - **Fluorescence in situ hybridization (FISH)** uses **fluorescently labeled DNA probes** that bind to specific complementary **DNA sequences** on chromosomes. - It is a powerful cytogenetic technique for detecting **chromosomal abnormalities**, such as deletions, translocations, and amplifications, thereby identifying abnormal gene sequences.
Question 6: Which type of mutation can act as a suppressor to restore the wild-type phenotype in organisms carrying a mutant gene?
- A. Frameshift mutation of coding gene
- B. Mutation of tRNA (Correct Answer)
- C. Deletion of mutant gene
- D. Addition of another normal gene
Explanation: ***Mutation of tRNA*** - A **tRNA suppressor mutation** can alter its anticodon, allowing it to recognize a **stop codon** (nonsense suppressor) or a missense codon, and insert an amino acid, thereby suppressing the original mutation. - This is a classic example of an **intergenic suppressor mutation** that acts at a different genetic locus from the original mutation. - These suppressors are particularly effective for **nonsense mutations** (premature stop codons) and certain missense mutations by correcting the decoding error during translation. *Frameshift mutation of coding gene* - A single frameshift mutation causes a shift in the **reading frame**, leading to a completely different protein sequence downstream and often a premature stop codon, which would worsen the phenotype. - While a **second compensating frameshift** mutation in the same gene could theoretically restore the reading frame (acting as an intragenic suppressor), this is context-dependent and less reliable than tRNA suppressors. - The question asks for mutations that "can act as a suppressor," and **tRNA mutations are the more universally recognized and reliable suppressor mechanism** in classical genetics. *Deletion of mutant gene* - **Deleting the mutant gene** removes the genetic information entirely but does not restore wild-type function; instead, it typically results in **loss of function** or complete absence of the protein. - This would lead to a **null phenotype** rather than restoration of wild-type phenotype, especially if the gene is essential. *Addition of another normal gene* - The **addition of another normal (wild-type) gene copy** provides a functional protein that can compensate for the mutant gene's deficiency. - While this can restore a wild-type phenotype, it represents **gene complementation** or gene therapy, not a true suppressor mutation that modifies the interpretation or expression of the existing mutant allele.
Question 7: Which malformation is associated with mutations in the HOX gene?
- A. Polysyndactyly (Correct Answer)
- B. Holoprosencephaly
- C. Mayer Rokitansky syndrome
- D. Gorlin syndrome
Explanation: ***Polysyndactyly*** - The **HOX gene** plays a critical role in limb development and is associated with the malformation of **polysyndactyly**, which is characterized by extra fingers or toes [1]. - This condition is due to the disruption of the normal **patterning** during limb formation, directly involving the action of HOX genes [1]. *Gorlin syndrome* - Gorlin syndrome is primarily caused by mutations in the **PTCH1 gene**, linked to **basal cell carcinoma** and other abnormalities. - It does not involve HOX gene mutations, hence is **not** related to limb malformations. *Holoprosencephaly* - Holoprosencephaly is a developmental condition often linked to **chromosomal anomalies** and abnormal embryonic development, **not specifically** HOX gene mutations. - It refers to the incomplete separation of the forebrain, distinct from the **limb malformations** associated with HOX genes. *Mayer Rokitansky syndrome* - Mayer-Rokitansky syndrome involves **agenesis** or **hypoplasia** of the uterus and upper two-thirds of the vagina, which is due to other genetic factors. - This condition is not related to the functions of the **HOX genes** in limb or skeletal development. **References:** [1] Kumar V, Abbas AK, et al.. Robbins and Cotran Pathologic Basis of Disease. 9th ed. Bones, Joints, and Soft Tissue Tumors, p. 1186.
Question 8: A 15-year-old boy returned from a vacation to his relatives' village. He developed a severe headache and purulent nasal discharge and was diagnosed with primary amebic meningoencephalitis (PAM). He died 5 days later. Which is the most likely etiological agent?
- A. Plasmodium falciparum
- B. Entamoeba histolytica
- C. Naegleria fowleri (Correct Answer)
- D. Toxoplasma
Explanation: ***Naegleria fowleri*** - This free-living amoeba is the causative agent of **Primary Amebic Meningoencephalitis (PAM)**, a rapidly fatal infection acquired through nasal exposure to contaminated warm freshwater. - The rapid progression from symptom onset (headache, purulent nasal discharge) to death within 5 days is characteristic of PAM caused by **_Naegleria fowleri_**. *Plasmodium falciparum* - This parasite causes **falciparum malaria**, which can lead to cerebral malaria, but its presentation involves cyclic fevers, chills, and typically a longer disease course and different diagnostic markers than PAM. - While it affects the brain, it does not cause purulent nasal discharge or the rapid, fulminant meningoencephalitis seen in PAM. *Entamoeba histolytica* - This amoeba causes **amebic dysentery** and, in rare cases, extraintestinal amebiasis like liver abscesses; it does not typically cause PAM. - Central nervous system involvement by _Entamoeba histolytica_ is usually secondary to systemic spread and manifests as abscesses, not a fulminant meningoencephalitis acquired via nasal passages. *Toxoplasma* - _Toxoplasma gondii_ causes toxoplasmosis, which can lead to toxoplasmic encephalitis, especially in immunocompromised individuals. - _Toxoplasma_ infection typically has a slower onset, different risk factors (e.g., raw meat, cat feces), and does not present with a rapid, purulent meningoencephalitis following swimming exposure.
Question 9: Transmission of R factor is by which mechanism?
- A. Conjugation (Correct Answer)
- B. Transduction
- C. Transformation
- D. Lysogenic conversion
Explanation: **Explanation:** **Why Conjugation is Correct:** The **R factor (Resistance factor)** is a type of plasmid that carries genes for antibiotic resistance. It consists of two components: the **Resistance Transfer Factor (RTF)**, which contains genes for autonomous replication and conjugative transfer, and the **r-determinant**, which carries the actual resistance genes. The primary mechanism for the spread of R factors between bacteria (especially Gram-negative bacilli like *E. coli* and *Salmonella*) is **Conjugation**. This process involves direct cell-to-cell contact via a sex pilus, allowing the rapid horizontal transfer of multi-drug resistance across different bacterial species. **Why Other Options are Incorrect:** * **Transduction:** This involves the transfer of DNA via a **bacteriophage** (virus). While some resistance genes (like those for Penicillinase in *Staphylococci*) can be transduced, the large R factor plasmid is typically transferred via conjugation. * **Transformation:** This is the uptake of **naked DNA** from the environment. It is a significant mechanism for species like *S. pneumoniae* and *Neisseria*, but it is not the classic route for R factor transmission. * **Lysogenic Conversion:** This occurs when a temperate phage integrates into the bacterial chromosome (prophage), imparting new phenotypic traits (e.g., **Diphtheria toxin**, Cholera toxin, or Erythrogenic toxin). It does not involve R factor transfer. **High-Yield Clinical Pearls for NEET-PG:** * **R Factor:** Responsible for "Infectious Drug Resistance." One R factor can carry resistance to multiple drugs (e.g., Sulfonamides, Streptomycin, Chloramphenicol). * **Conjugation:** The most common method for the spread of multidrug resistance in clinical settings. * **Transposons ("Jumping Genes"):** These are DNA sequences that can move from a plasmid to a chromosome (or vice versa) and are often found within R factors.
Question 10: Resistance to drugs in tuberculosis develops by which mechanism?
- A. Transduction
- B. Conjugation
- C. Transformation
- D. Mutation (Correct Answer)
Explanation: **Explanation:** In *Mycobacterium tuberculosis* (MTB), drug resistance is primarily driven by **spontaneous genetic mutations** in specific chromosomal genes. Unlike many other bacteria, MTB does not possess horizontal gene transfer mechanisms like plasmids or transposons. **1. Why Mutation is Correct:** Resistance in MTB occurs due to random, stepwise mutations in the chromosomal DNA. When a patient is treated with inadequate monotherapy or irregular dosing, these resistant mutants are "selected" and multiply (selective pressure). * **High-yield examples:** * **Isoniazid (INH) resistance:** Mutations in the *katG* gene (loss of catalase-peroxidase activity) or *inhA* gene. * **Rifampicin resistance:** Mutations in the *rpoB* gene (beta-subunit of RNA polymerase). **2. Why Other Options are Incorrect:** * **Transduction (A):** Involves DNA transfer via a bacteriophage. While mycobacteriophages exist, they do not play a role in clinical drug resistance. * **Conjugation (B):** Involves cell-to-cell contact via a sex pilus to transfer plasmids. MTB lacks the plasmids necessary for this process. * **Transformation (C):** Involves the uptake of free "naked" DNA from the environment. This is not a documented mechanism for resistance in MTB. **Clinical Pearls for NEET-PG:** * **Multidrug-Resistant TB (MDR-TB):** Defined as resistance to at least **Isoniazid and Rifampicin**. * **Extensively Drug-Resistant TB (XDR-TB):** MDR-TB plus resistance to any **fluoroquinolone** and at least one **Group A drug** (Bedaquiline or Linezolid). * **Genotypic Testing:** Molecular methods like **GeneXpert (CBNAAT)** and **Line Probe Assay (LPA)** detect resistance by identifying these specific chromosomal mutations (e.g., *rpoB* for Rifampicin).
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