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NEET: PCR Made Easy With Solutions, Components, Principle, Steps, PCR Cycle, Applications

NEET: PCR Made Easy With Solutions, Components, Principle, Steps, PCR Cycle, Applications

Edited By Irshad Anwar | Updated on Jun 04, 2022 09:49 AM IST | #NEET
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How Important Is PCR For NEET?

The topic PCR is from the chapter Biotechnology: Principles and Processes. According to the analysis of the past five years' NEET papers, 20 questions were asked from the chapter - Biotechnology: Principles and Processes, which has approximately 9% weightage of the total questions asked from Zoology. Out of 20, five questions were based on the PCR concept alone. You will find those questions from PCR at the end of the article. Let's understand the PCR in detail.

NEET: PCR Made Easy With Solutions, Components, Principle, Steps, PCR Cycle, Applications
NEET: PCR Made Easy With Solutions, Components, Principle, Steps, PCR Cycle, Applications

Introduction

The polymerase chain reaction, commonly referred to as PCR, is a technique used to create multiple copies of a certain segment of DNA in vitro. Kary Mullis, an American biochemist, developed this technique in 1983. PCR technique is used to generate millions of copies of a DNA segment. It allows the scientist to amplify a very small DNA sample to a large amount for a detailed study. PCR is usually used in biotechnology labs, several areas of biology and in medicine such as medical diagnostics, molecular biology research and even in some areas of ecology. The goal of PCR is to produce amplified DNA so that it can be analysed or used.

Background wave

Components Of PCR

  • DNA Template - The DNA of interest to be amplified.

  • Taq Polymerase - It is a DNA polymerase I. PCR requires the enzyme DNA polymerase to synthesise new strands of DNA. The DNA polymerase used in PCR is known as Taq polymerase. As it is isolated from the heat-tolerant bacterium Thermus aquaticus, this enzyme can withstand high temperatures.

T. aquaticus resides in hot water springs and its DNA polymerase is heat stable and remains most active at about 70 degrees celsius - the temperature at which DNA polymerase of E. coli and humans would be non-functional. As the high temperature is repeatedly used in PCR to denature the DNA strands, the Taq polymerase is considered ideal for PCR due to its high heat stability.

  • Primers -

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1653636758489

They are short chemically synthesised oligonucleotides, single-stranded DNA complementary to that particular segment of DNA to be amplified. The 2 primers that are used in each PCR are designed in a way to flank the target region of the DNA - region to be copied or amplified. The primers bind to the template strand via complementary base pairing. After binding they can be extended due to the action of DNA polymerase.

  • dNTPs - It is deoxyribonucleotide triphosphate, the building block to synthesise DNA. The energy required for the polymerisation is provided by them as well. The four common dNTPs used in PCR are adenine (dATP), thymine (dTTP), cytosine (dCTP), and guanine (dGTP).

  • Buffer - Potassium and Magnesium present in the buffer solution provide optimum conditions for denaturation and renaturation of DNA. Buffers also play an important role in polymerase activity, stability, and fidelity.

Principle Of PCR

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The technique is based on the replication of DNA using enzymes and majorly relies on thermal cycling. A particular segment of DNA is amplified in vitro - in an artificial environment outside the body - to make new strands of DNA using the existing strand as a template, enzyme Taq DNA polymerase and two sets of primers. This particular segment of DNA to be amplified can be a gene of interest for a researcher, a genetic marker used in forensic science to match the DNA from the crime scene with that of the suspects or a gene whose function is to be determined by the experimenter.

What are the steps in correct order involved in a polymerase chain reaction?

The Steps Of PCR

The key ingredients used in polymerase chain reaction are template DNA, primers, DNA polymerase i.e. Taq Polymerase, and nucleotides - the DNA building blocks. All the ingredients are kept in a tube along with the co-factors required by the enzyme. Then the tube goes through the repeated cycle of heating and cooling and thus new strands of DNA are synthesised from the template DNA.

What has happened inside the tube due to repeated heating and cooling?

Each cycle of PCR includes three steps:- 

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1653636759370

Step 1: Denaturation

The first step is denaturation in which double-stranded DNA is separated by heating the sample. It takes place at about 94 degrees Celsius for about 30 seconds. At this temperature, the hydrogen bonds between the base pairs are broken and thus single-stranded DNA is obtained for the next step.

Step 2: Annealing

In the second step of PCR, the temperature is lowered up to about 54 to 64 degrees celsius. This cooling allows primers to bind their complementary sequences on the template strand. The two primers - a forward primer and a reverse primer - bind to the 2 separated template strands running in opposite directions. After complementary base pairing, DNA polymerase binds at primer-template hybrid and initiates the DNA synthesis. Thus, primers act as initiating points for DNA synthesis.

Step 3: Extension or Elongation

In this step, the temperature is again raised to 72 degrees celsius. For elongation or extension, a thermostable DNA polymerase is used - Taq polymerase. Taq Polymerase extends the primers by adding the nucleotides - 1000 base pairs per minute under the optimum conditions - in the 5’ to 3’ direction and thus the complementary strand of the DNA template is synthesised. As a result of 1 cycle of PCR, a double-stranded molecule is obtained in which one is a newly synthesised strand and another is an original strand.

PCR Cycle

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This cycle of 3 steps is repeated many times to get billions of copies in a very short period. In a typical PCR reaction, the cycle is repeated 25–35 times and it usually takes 2-4 hours. However, the time taken depends on the number of cycles and the length of the DNA region to be copied. The results of PCR can be visualised using gel electrophoresis.

The original DNA segment is not used as a template each time. The newly formed DNA in one round can serve as a template in the next round. Numerous molecules of Taq polymerase and copies of primers float around the reaction. So, molecules of DNA roughly double in each round of cycling as shown in the diagram above.

Applications Of PCR/Reasons To Perform PCR

PCR is applied in many research labs as an essential tool in research and biotechnology. It has numerous practical applications in diagnostics, forensics, and genetic testing. PCR is a susceptible technique and provides results rapidly. The desired gene can be turned into millions and billions of copies that can be utilised for several purposes such as analysis, cloning and sequencing. Some of the essential applications in various fields are given below:

Diagnostics

  • PCR can be used to diagnose the presence of a viral or bacterial genome in the host cell even before the onset of the disease. For example, the viral load can be detected using the PCR technique: RT-PCR is used to detect the presence of the SARS-CoV-2 viral genome.

  • To test the genetic disease mutation such as oncogene mutation. Early detection helps in the treatment of various types of cancer such as lymphoma and leukaemia.
  • To monitor the gene in gene therapy.
  • To detect the disease-causing genes in the parents.

Genetics And Research

  • To compare the genome of two organisms in genomic studies

  • To study gene expression

  • Gene Mapping

  • To create the mutant genes to study site-directed mutagenesis

Forensics

  • Used as a tool in genetic fingerprinting.

  • Identifying the criminal from the suspects- A small DNA sample collected from the crime site can be amplified to analyse and compare with that of the suspects

  • Paternity tests

Recombinant DNA Technology

  • The gene of interest is amplified using PCR and then inserted into a vector for cloning.

Fossils

  • Ancient DNA samples can be detected and analysed using PCR


Recommended Video

Important Questions

Question 1: What is PCR?

Answer: It is a technique used to create multiple copies of a certain segment of DNA in vitro. Kary Mullis, an American biochemist, developed this technique in 1983. Generating millions of copies of a DNA segment has been possible because of the PCR technique. It allows the scientist to amplify a very small DNA sample to a large amount for a detailed study.

Question 2: The correct order of steps in Polymerase Chain Reaction (PCR) is________. (NEET 2018)

Answer: The correct order in which polymerase chain reaction takes place is:

  1. Denaturation

  2. Annealing

  3. Elongation or Extension

Question 3. What is the importance of PCR?

Answer: PCR is applied in many research labs as an essential tool in research and biotechnology. It has numerous practical applications in diagnostics, forensics, and genetic testing. PCR is a susceptible technique and provides results rapidly. The desired gene can be turned into millions and billions of copies that can be utilised for several purposes such as analysis, cloning and sequencing.

Question 4: During the process of gene amplification using PCR, if a very high temperature is not maintained in the beginning, then which step of PCR will be affected first? (NEET 2021)

Answer: Denaturation

In denaturation, the double-stranded DNA is separated by heating the sample. It takes place at about 94 degrees celsius for about 30 seconds. At this temperature, the hydrogen bonds between the base pairs are broken and thus single-stranded DNA is obtained for the next step.

Question 5: Which of the following is not an application of PCR (Polymerase Chain Reactionando)? (NEET 2021)

(A) Molecular diagnosis

(B) Gene amplification,

(C) Purification of the isolated protein,

(D) Detection of gene mutation.

Answer: (C) Purification of the isolated protein.

Some important areas of application of PCR are given below:

(i) Diagnosis of pathogens

(ii) Diagnosis of a specific mutation.

(iii) DNA fingerprinting

(iv) Detection of specific microorganisms

(v) In prenatal diagnosis

(vi) Diagnosis of plant pathogens

(vii) In palaeontology

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Many websites I have seen written the cutoff for NEET 2018 West Bengal state quota was around 360 for SC category... Is it true?

Hello Rahul,

yes, the NEET 2018 cutoff fot the SC category in west  bangal was around 360 marks for the state quota. however the exact cutoff may vary slightly depending on factors like the number of applicants and seat availability. its always good to refer to the official counselling details for precise information.

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Riya Dhoot 01 April,2025
How to get admission in bams college without neet

Hello dear,

To get admission to a BAMS course without NEET, here are a few possible routes:

  1. State-Level Exams: Some states, like Karnataka, Maharashtra, or Uttarakhand, may conduct their own entrance exams for BAMS admissions. These may not require NEET scores.

  2. Private Colleges & Management/NRI Quota: Some private colleges may offer admission through management or NRI quotas without requiring NEET scores, although the fees may be higher.

  3. Special Exemptions: Rarely, some colleges may offer exemptions or internal entrance exams, but this is not common.

Make sure the college is recognized by (CCIM) to ensure the degree is valid. Always verify the latest admission guidelines from the specific state or college.

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Devanshi Goyal 01 April,2025
How to get admission in bams college without neet

You cannot get admission in BAMS college without neet because it is compulsory for every medical student to give NEET (entrance exam) for the admission in any medical college. You have to qualify the exam for the admission if you want to join private college

Read Complete Answer
Harsha Prajapat 01 April,2025
Make suitable schedule for neet 2025 with In 10 days

The key is to focus on revision, important topics, and time management. Here’s a tailored 10-day plan:

Day 1-2: Focus on Physics (Core Concepts & Formulas)

  • Morning (3 hours):

    • Revise Mechanics: Laws of Motion, Work-Energy, Rotational Motion.

    • Focus on Electrostatics: Coulomb’s Law, Electric Field, Potential.

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    • Solve MCQs from Mechanics & Electrostatics.

    • Focus on time-based questions.

  • Evening (2 hours):

    • Revise Important formulas in Physics.

    • Short-break activities: Quick review videos or notes.

  • Night (1 hour):

    • Practice NEET Previous Year Questions (Physics).

Day 3-4: Focus on Chemistry (Inorganic & Organic)

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    • Focus on Organic Chemistry: General Organic Chemistry, Hydrocarbons, Alcohols, and Ethers.

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    • Solve MCQs from Inorganic and Organic Chemistry.

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    • Quick revisions of key concepts.

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Day 5-6: Focus on Biology (Botany)

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Day 9: Revision Day

  • Morning (3 hours):

    • Revise Physics concepts: Quick review of important chapters.

    • Practice Physics MCQs on weak topics.

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    • Revise Chemistry: Quick concept review and focus on problem-solving.

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    • Do quick practice tests.

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Day 10: Full Mock Test & Final Revision

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    • Focus on short notes and key formulas.

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    • Mental Relaxation: Avoid heavy study. Do some light revision or relaxation activities. Rest well for the exam.

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Devanshi Goyal 01 April,2025
In the month may 2025, I will get my 12 result .. So I want government college for bsc in microbiology.so suggest some ..I had not given neet and cuet ..so without that suggest me ..

Dear

If you're looking to pursue a BSc in Microbiology from a government college without appearing for NEET or CUET, you can still explore many excellent options. Most government colleges admit students to BSc courses based on the marks obtained in your 12th-grade exams. Here are some of the top government colleges you can consider for BSc in Microbiology:

1. University of Delhi (DU)

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  • Admission Process: Based on 12th-grade marks and cutoffs, which vary every year.
  • Eligibility: You need to have Biology as a subject in Class 12 and meet the cutoff requirements.

2. Banaras Hindu University (BHU)

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  • Admission Process: Admission is usually based on the entrance test, but if you’re looking for colleges with direct 12th marks-based admission, consider other options.
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3. University of Calcutta

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  • Admission Process: Based on merit (your 12th-grade marks).
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4. Jamia Millia Islamia (JMI)

  • Courses: BSc in Microbiology.
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5. Madras University

  • Courses: BSc in Microbiology.
  • Admission Process: Based on merit, considering your marks in 12th.
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6. University of Pune (Savitribai Phule Pune University)

  • Courses: BSc in Microbiology.
  • Admission Process: Based on 12th-grade marks.
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7. Karnataka University, Dharwad

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8. St. Xavier’s College, Mumbai

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9. Aligarh Muslim University (AMU)

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10. Sri Venkateswara College, Delhi University

  • Courses: BSc (Hons) Microbiology.
  • Admission Process: Based on Class 12 marks through the DU admission process.
  • Eligibility: Class 12 with Biology as a subject.

Tips:

  • Research and Apply Early: Keep an eye on the official websites for admission dates and specific criteria.
  • Check Cutoffs: Colleges like DU and BHU release cutoff lists, which are based on your 12th-grade performance. Ensure you meet the required percentage.
  • Prepare Documents: Make sure to have your Class 12 marksheet and necessary documents ready for the application.

These colleges offer great opportunities for pursuing BSc in Microbiology, and you do not need to appear for NEET or CUET for admission to these programs.

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NEET MBBS cut-off rises for Bangalore Medical College, drops for other Karnataka govt colleges
April 02, 2025 - 04:59 PM IST
Aiming for 650+ in NEET 2025? Focus on these high-weightage chapters
April 01, 2025 - 11:55 AM IST
Sri Chaitanya Academy, Infinity Learn launch NEET preparation course, 'ATREYA'
March 29, 2025 - 10:13 PM IST
Lowest NEET cut-off for MBBS in Maharashtra government medical colleges; passing mark out of 720
March 26, 2025 - 09:34 AM IST
NEET aspirant from Bihar dies by suicide in Kota hostel; ninth incident this year
March 26, 2025 - 07:17 AM IST
Medical courses after Class 12th without NEET 2025; course-wise eligibility, career prospectus
March 21, 2025 - 08:38 PM IST
Aakash launches free rapid revision course for NEET; paid crash course for JEE Advanced
March 21, 2025 - 03:32 PM IST

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