Biochemical Techniques Practice Questions

Q: Which of the following techniques is used for the detection of variation in DNA sequence and gene expression?

Microarray. **Explanation:** The correct answer is **Microarray**. This technique is uniquely capable of analyzing thousands of genes simultaneously. It utilizes a solid surface (chip) containing microscopic spots of DNA probes. When labeled sample DNA or cDNA binds to these probes, it allows for the detection of **Single Nucleotide Polymorphisms (SNPs)**—representing variations in DNA sequence—and the quantification of mRNA levels, which reflects **gene expression**. **Analysis of Options:** * **A. Western blot:** Used specifically for the detection and quantification of **proteins** using antibody-antigen interactions. It does not analyze DNA or RNA. * **B. Northern blot:** Used for the detection of specific **RNA** sequences to study gene expression. However, it cannot detect DNA sequence variations (mutations/polymorphisms) and typically analyzes one gene at a time. * **C. Southern blot:** Used for the detection of specific **DNA** sequences (e.g., RFLP). While it detects DNA variations, it cannot measure gene expression (mRNA). * **D. Microarray:** The "high-throughput" nature of microarrays allows for the simultaneous assessment of both structural variations (DNA) and functional activity (Expression). **High-Yield Clinical Pearls for NEET-PG:** * **Mnemonic (SNoW DRoP):** **S**outhern-**D**NA; **N**orthern-**R**NA; **W**estern-**P**rotein. * **Southwestern Blot:** Used to detect **DNA-binding proteins** (e.g., transcription factors like c-Jun or c-Fos). * **Clinical Use of Microarray:** Widely used in oncology (to categorize tumors based on gene expression profiles) and in prenatal testing to detect submicroscopic chromosomal deletions or duplications.

Q: In which type of chromatography is the separation of protein molecules based on their size, shape, and molecular weight?

Gel filtration chromatography. ### Explanation **Correct Answer: B. Gel filtration chromatography** **Why it is correct:** Gel filtration chromatography, also known as **Size-Exclusion Chromatography (SEC)** or **Molecular Sieving**, separates proteins based on their **size, shape, and molecular weight**. The stationary phase consists of porous beads (e.g., Sephadex, agarose). * **Mechanism:** Small molecules enter the pores of the beads, increasing their path length and slowing their movement. Large molecules are "excluded" from the pores and travel only through the spaces between beads. * **Result:** Large molecules elute **first**, while smaller molecules elute **later**. **Why the other options are incorrect:** * **A. Partition chromatography:** Separation is based on the differential solubility of solutes between two phases (liquid-liquid or liquid-gas), such as in Paper Chromatography. * **C. Ion-exchange chromatography:** Separation is based on the **net surface charge** of the proteins. It uses a charged stationary phase (anion or cation exchangers) to bind proteins of the opposite charge. * **D. Affinity chromatography:** Separation is based on **high-specificity biological interactions**, such as enzyme-substrate, antigen-antibody, or hormone-receptor binding. It is the most specific method for protein purification. **High-Yield Clinical Pearls for NEET-PG:** * **Elution Order:** In Gel Filtration, the molecule with the **highest molecular weight** elutes first (Void Volume). * **Desalting:** Gel filtration is commonly used to remove small salt molecules from a protein solution. * **Determining MW:** It is a standard technique used to estimate the quaternary structure and molecular weight of native proteins. * **Common Matrices:** Sephadex (Dextran), Sepharose (Agarose), and Bio-Gel (Polyacrylamide).

Q: Which preservative is used for blood collection?

Sodium fluoride. **Explanation:** **Sodium fluoride (NaF)** is the preferred preservative for blood glucose estimation. Its primary role is to inhibit **enolase**, a key enzyme in the glycolytic pathway. Without a preservative, red blood cells continue to metabolize glucose *in vitro*, leading to a decrease in glucose levels by approximately 5–7% per hour. By blocking glycolysis, NaF ensures that the glucose concentration remains stable for up to 48–72 hours. **Analysis of Options:** * **Sodium fluoride (Correct):** Acts as an antiglycolytic agent. It is typically used in combination with potassium oxalate (the anticoagulant) in **grey-topped tubes**. * **Thymol (Incorrect):** This is a preservative used for **24-hour urine collections**, primarily to inhibit bacterial growth and preserve chemical constituents, not for blood collection. * **Potassium oxalate (Incorrect):** While often present in the same tube as NaF, it is an **anticoagulant**, not a preservative. It prevents clotting by precipitating calcium ions. * **No preservative (Incorrect):** If no preservative is used (e.g., plain red-top tube), glycolysis continues, leading to falsely low glucose readings (pseudohypoglycemia). **High-Yield Clinical Pearls for NEET-PG:** * **Mechanism:** NaF inhibits enolase by forming a complex with magnesium and phosphate. * **Interference:** NaF should **not** be used for samples intended for **Urease-based urea estimation**, as fluoride inhibits the urease enzyme. * **Electrolytes:** NaF can interfere with certain electrolyte assays and enzyme studies (like ALP or Amylase) due to its chelating properties. * **Ratio:** The standard mixture in grey tubes is 1 part NaF to 3 parts Potassium Oxalate.

Q: DNA restriction is done by which of the following methods?

Agarose gel electrophoresis. **Explanation:** **Why Agarose Gel Electrophoresis is correct:** DNA restriction involves cutting DNA into fragments using restriction endonucleases. Once cut, these fragments must be separated and analyzed based on their size. **Agarose gel electrophoresis** is the standard technique used for this purpose. DNA molecules are negatively charged (due to the phosphate backbone) and migrate toward the positive electrode (anode) when placed in an electric field. The agarose matrix acts as a molecular sieve; smaller DNA fragments move faster and further through the pores than larger ones, allowing for precise separation and visualization (usually with Ethidium Bromide). **Why other options are incorrect:** * **Paper Chromatography:** This technique is primarily used to separate small soluble molecules like amino acids or sugars based on their solubility and partition coefficients, not large macromolecules like DNA. * **Spectrophotometry:** This is used to **quantify** the concentration and purity of DNA (measuring absorbance at 260 nm and 280 nm) but cannot separate or "restrict" DNA fragments by size. * **Spectrometry (e.g., Mass Spectrometry):** While used for identifying the chemical composition or mass of small molecules and proteins, it is not the standard method for routine DNA restriction analysis in a clinical or research lab. **High-Yield Clinical Pearls for NEET-PG:** * **DNA Charge:** DNA is polyanionic; it always moves from **Negative (Cathode) to Positive (Anode)**. * **Visualization:** **Ethidium Bromide (EtBr)** is the most common intercalating agent used to visualize DNA under UV light. * **Purity Check:** A **260/280 ratio of ~1.8** indicates pure DNA. A lower ratio suggests protein contamination. * **Pulsed-Field Gel Electrophoresis (PFGE):** A variation used to separate extremely large DNA fragments (e.g., whole chromosomes).

Q: What does a microarray primarily study?

Multiple genes. **Explanation:** **Microarray technology** is a high-throughput biochemical technique used to analyze the expression levels of thousands of genes simultaneously. It utilizes a solid surface (usually a glass slide or silicon chip) onto which microscopic spots of DNA probes are attached. When a sample of fluorescently labeled cDNA or cRNA is applied, it hybridizes with the complementary probes, allowing researchers to determine which genes are "turned on" or "off" in a specific tissue or under specific conditions. * **Why Option A is correct:** Microarrays are specifically designed for **transcriptomics** and **genomics**. They allow for the simultaneous study of the entire transcriptome (multiple genes) rather than focusing on a single gene (as seen in Northern blotting). * **Why Option B is incorrect:** While microarrays can be used to *diagnose* or research diseases (e.g., cancer subtyping), they primarily study the **genetic expression** underlying the disease, not the disease entity itself. * **Why Option C is incorrect:** Microarrays study molecular components (DNA/RNA) within cells; they do not study whole organisms. * **Why Option D is incorrect:** Blood grouping is typically performed via serological agglutination tests or specific PCR for genotypes, not broad-scale microarray analysis. **High-Yield NEET-PG Pearls:** 1. **Southern Blot:** Detects DNA. 2. **Northern Blot:** Detects RNA. 3. **Western Blot:** Detects Proteins. 4. **Microarray:** Often considered a "multiplex" version of Northern/Southern blotting. 5. **Clinical Use:** Microarrays are the "gold standard" for detecting **Copy Number Variations (CNVs)** in children with developmental delays or congenital anomalies (Chromosomal Microarray).

Q: Karyotyping under light microscopy is done by which banding technique?

G banding. **Explanation:** **G-banding (Giemsa banding)** is the most common and standard technique used for routine karyotyping under light microscopy. In this method, chromosomes are first treated with **Trypsin** (to partially digest proteins) and then stained with **Giemsa stain**. This produces a characteristic pattern of alternating light and dark bands: * **Dark bands (G-positive):** Represent AT-rich, gene-poor, heterochromatic regions that replicate late. * **Light bands (G-negative):** Represent GC-rich, gene-dense, euchromatic regions that replicate early. **Analysis of Incorrect Options:** * **R-banding (Reverse):** This is the "reverse" of G-banding. Chromosomes are heat-denatured before staining. It is used specifically to study the distal ends (telomeres) of chromosomes. * **Q-banding (Quinacrine):** This was the first banding method developed. It uses quinacrine mustard (a fluorescent stain) and requires a **fluorescence microscope**, not a standard light microscope. * **C-banding (Constitutive Heterochromatin):** This technique specifically stains the centromeres and regions containing constitutive heterochromatin (like chromosomes 1, 9, 16, and Y). **High-Yield Clinical Pearls for NEET-PG:** * **Sample Collection:** For postnatal karyotyping, **Peripheral Blood Lymphocytes** are used. They must be stimulated with a mitogen like **Phytohemagglutinin (PHA)**. * **Cell Cycle Arrest:** Cells are arrested in **Metaphase** using **Colchicine** (which inhibits spindle formation) because chromosomes are most condensed and visible during this stage. * **Resolution:** Standard G-banding identifies roughly 400–550 bands per haploid set, allowing for the detection of numerical and large structural abnormalities.

Question 1

Which of the following techniques is used for the detection of variation in DNA sequence and gene expression?

Accepted Answer

Microarray

Answer

Western blot

Answer

Northern blot

Answer

Southern blot

Question 2

In which type of chromatography is the separation of protein molecules based on their size, shape, and molecular weight?

Accepted Answer

Gel filtration chromatography

Answer

Partition chromatography

Answer

Ion-exchange chromatography

Answer

Affinity chromatography

Question 3

Which preservative is used for blood collection?

Accepted Answer

Sodium fluoride

Answer

Thymol

Answer

Potassium oxalate

Answer

No preservative is needed

Question 4

Nephelometry is a technique used in the measurement of:

Accepted Answer

Immunoglobulins

Answer

Refraction of light

Answer

Size of renal stones

Answer

Optical density of fluids

Question 5

DNA restriction is done by which of the following methods?

Accepted Answer

Agarose gel electrophoresis

Answer

Paper chromatography

Answer

Spectrophotometry

Answer

Spectrometry

Question 6

Which substance can be confused with protein during a biochemical estimation test?

Accepted Answer

Phosphates

Answer

Nitrates

Answer

Sulphates

Answer

Bile salts

Question 7

Reliability is tested by which of the following?

Accepted Answer

R-chart

Answer

Mean chart

Answer

Levy Jennings chart

Answer

Shewhart control chart

Question 8

What does a microarray primarily study?

Accepted Answer

Multiple genes

Answer

Diseases

Answer

Organisms

Answer

Blood groups

Question 9

Which of the following is the strongest type of chemical bond?

Accepted Answer

Electrostatic bond

Answer

Hydrogen bond

Answer

Hydrophobic interaction

Answer

Van der Waals force

Question 10

Karyotyping under light microscopy is done by which banding technique?

Accepted Answer

G banding

Answer

R banding

Answer

Q banding

Answer

C banding

Biochemical Techniques — MCQs

Biochemical Techniques — MCQs

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