Degenerate primers in PCR
Which of the following statements about degenerate primers is not correct?
a. Because of the nature of degenerate primers, the annealing temperature during PCR using these primers must be lowered to account for the mismatches.
b. Degenerate primers are often designed by working backwards from a known amino acid sequence.
c. Degenerate primers are used even when the sequence of DNA is known.
d. Within a population of degenerate primers, some will bind perfectly, some will bind with mismatches, and others will not bind.
Correct Answer: (c) Degenerate primers are used even when the sequence of DNA is known.
Explanation
Degenerate primers are a mixture of similar but not identical primers designed to account for variability in the DNA sequence. They are particularly useful when the exact DNA sequence is unknown or when working from a known protein sequence. The goal is to increase the chances of successful amplification by covering multiple possible codon combinations that encode the same amino acid sequence.
1. What Are Degenerate Primers?
- Degenerate primers are designed when the exact nucleotide sequence is unknown but the amino acid sequence is known.
- Because the genetic code is redundant (multiple codons code for the same amino acid), degenerate primers are designed to cover all possible codon combinations.
- They are particularly useful in amplifying genes from different species or homologous genes with sequence variability.
2. Why the Other Statements Are True
| Statement | Explanation | True/False |
|---|---|---|
| (a) Because of the nature of degenerate primers, the annealing temperature during PCR using these primers must be lowered to account for the mismatches. | Degenerate primers have sequence mismatches, which reduce binding efficiency. Lower annealing temperature improves primer-template binding. | ✅ True |
| (b) Degenerate primers are often designed by working backwards from a known amino acid sequence. | Degenerate primers are created by considering the codon degeneracy of the known protein sequence. | ✅ True |
| (c) Degenerate primers are used even when the sequence of DNA is known. | If the DNA sequence is known, specific primers (not degenerate primers) are preferred for higher specificity and efficiency. | ❌ False |
| (d) Within a population of degenerate primers, some will bind perfectly, some will bind with mismatches, and others will not bind. | Degenerate primers are a mixture; some will match better than others. | ✅ True |
3. Why Option (c) Is Incorrect
- If the DNA sequence is already known, specific primers are used instead of degenerate primers because:
- Specific primers improve efficiency and accuracy.
- Degenerate primers are used only when the exact DNA sequence is unknown or when there is sequence variability.
- Therefore, using degenerate primers when the sequence is known is unnecessary and inefficient.
4. How Degenerate Primers Are Designed
1. Start with a Protein Sequence:
- Use a known protein sequence to predict possible codon combinations.
2. Design Multiple Primers:
- Create primers that cover all possible codon variations due to codon degeneracy.
3. Optimize Primer Binding:
- Lower annealing temperature to allow mismatched binding.
🚀 5. Applications of Degenerate Primers
1. Gene Cloning:
- Used when the target gene sequence is unknown but a similar gene sequence from another species is available.
2. Evolutionary Studies:
- Used to amplify homologous genes in different species.
3. Diagnostic Tests:
- Detecting viruses and pathogens with high sequence variability.
4. Protein Engineering:
- Identifying genes encoding proteins with similar functional domains.
6. Challenges with Degenerate Primers
1. Low Specificity:
- Mismatched primers can lead to non-specific amplification.
2. Reduced Efficiency:
- Mixed primer binding lowers reaction efficiency.
3. Complex Analysis:
- Multiple PCR products require further validation and sequencing.
7. Strategies to Improve Degenerate Primer Performance
1. Lower Annealing Temperature: Increases primer binding efficiency.
2. Use Nested PCR: Improves specificity by amplifying a smaller, internal region.
3. Optimize Mg²⁺ Concentration: Enhances polymerase activity and primer binding.
Summary
- Degenerate primers are used when the exact DNA sequence is unknown but the protein sequence is known.
- They are designed to account for codon variability and improve chances of successful amplification.
- If the sequence is known, specific primers should be used for greater accuracy and efficiency.
- Correct Answer: (c) Degenerate primers are used even when the sequence of DNA is known.
60 Comments
Akshay mahawar
March 17, 2025Done 👍
Ujjwal
March 17, 2025Okh sir
Suman bhakar
March 17, 2025Ok sir
Nisha
March 17, 2025Done
Arushi
March 17, 2025👍👍
Parul
March 22, 2025Done sir.
Soniya Shekhawat
August 24, 2025Degenerated primer is a random primer which is used in PCR when the DNA sequence is unknown.
Khushi Pareek
August 24, 2025If sequence is known, specific primers are designed
Mohd juber Ali
August 24, 2025Option c
In Degenrate primer nt sequence is unknown but amino acid sequence is known if dna sequence is already known then we use specific primer
Sakshi yadav
August 24, 2025Degenrate primers used even when the sequence of DNA polymerase is unknown not for known
Arushi Saini
August 24, 2025Specific primers are used if the DNA sequence is known
Avni
August 24, 2025Degenerate primers are designed when the exact nucleotide sequence is unknown but the amino acid sequence is known. If the DNA sequence is already known the specific primers are used instead of degenerate primers
Neha Yadav
August 24, 2025Degenerate primer are used even when the sequence of DNA is known is wrong statement because it is a random primer which is used when the DNA sequence is unknown and protein sequences is known
Roopal Sharma
August 24, 2025Degenrate primers are used when sequence of DNA is unknown
karishma don
August 24, 2025degenerate primers are used when the seq is not known, not when the sequence is known.
Khushi Agarwal
August 24, 2025Option c is false
Degenerate primers are used even when the sequence of DNA is known.
Anurag Giri
August 24, 2025Option c
Bcoz Degenerate primers are used when the exact DNA sequence is unknown but the protein sequence is known.
Aafreen Khan
August 24, 2025Degenerate primers are used even when the sequence of DNA is known
Dharmpal Swami
August 24, 2025Degenerate primer are use for unknown sequence of dna
Specific primer are use for known dna sequence
Aakanksha Sharma
August 24, 2025Option C is wrong bcz degenerate primer means random primer they are used when sequence is unknown
Priyanka Choudhary
August 24, 2025Degenerate primers are used even when the sequence of DNA is known
Dipti Sharma
August 24, 2025Degenerate primers are used when the sequence of DNA is unknown and amino acid sequence is known so option c is incorrect.
Karishma
August 24, 2025Done
Priya dhakad
August 24, 2025Degenerate primers are used even when the sequence of DNA is known.
AKANKSHA RAJPUT
August 24, 2025Option c
anjani sharma
August 24, 2025Option c is incorrect
Degenerate primers are used even when the sequence of DNA is known.
Sneha Kumawat
August 24, 2025Degenerate primers are random primer
When nucleotide sequence is unknown degenerate primers are used
Heena Mahlawat
August 24, 2025Degenerate primers are only used when sequence isn’t known
MOHIT AKHAND
August 24, 2025Done sir ✅
Santosh Saini
August 24, 2025Degenerate primers are used even when the sequence of DNA is unknown, so statement c is not correct
Ayush Dubey
August 24, 2025Degenerate primers are used even when the sequence of DNA is known.
Neelam Sharma
August 24, 2025Degenerate primers are used even when the sequence of DNA is known.
Aman Choudhary
August 24, 2025Degenerate primers are used when the sequence is unknown
Monika jangid
August 24, 2025Degenerated primer are random they use in unknown seq
Divya rani
August 25, 2025C is wrong because degenerate primers are used when dna sequence is unknown and when the Protein sequencing is used to known the unknown dna sequence.
shruti sharma
August 25, 2025Denature primer are used when the sequence is unknown
Payal Gaur
August 25, 2025Degenerate Primer are used for unknown sequence and specific primer for specific sequence/known sequence.
Bhavana kankhedia
August 26, 2025Degenerate primers are used even when the sequence of DNA is known.
Surendra Doodi
August 26, 2025Degenerate primer is used when the DNA sequence is unknown and protein sequences is known
Vanshika Sharma
August 26, 2025Degenerate primers are used in unkonw seq so ans 3 is correct
Manisha
August 26, 2025Degenerate primers are used when the sequence of DNA is unknown and amino acid sequence
Minal Sethi
August 26, 2025when DNA sequence is known then specific primers are used instead of degenerate primers
Shivani
August 26, 2025Option C is incorrect because Degenerate primers are used when the sequence of DNA is unknown and amino acid sequence is known.
Rishita
August 26, 2025Done
Pallavi Ghangas
August 26, 2025Degenerate primers are not used when sequence is known in PCR
Devika
August 26, 2025Option C.
Simran Saini
August 26, 2025Degenerate primers are used even when the sequence of DNA is known.
Mitali saini
August 28, 2025Correct Answer: (c) Degenerate primers are used even when the sequence of DNA is known.
Muskan singodiya
August 28, 2025Option c
Deepika Sheoran
August 28, 2025Option C is correct
Degenarate Primers used Even when the sequence of DNA polymerase is unknown not for known. Specific primer are use the known DNA sequence..
Neeraj Sharma
August 28, 2025If the sequence of the DNA is known then specific primers are designed
Parul Yadav
August 29, 2025Degenerate Primers are used when the sequence of the target DNA is not known but the Protein sequence is known.
Asha Gurzzar
August 30, 2025C statement is incorrect because degenerate primers are not used for known sequence of dna
Khushi Singh
August 30, 2025Degenerate primer are used when sequence is unknown
Kirti Agarwal
August 31, 2025Degenerate primers are used even when the sequence of DNA is known
Kajal
August 31, 2025Correct Answer is (c) Degenerate primers are used even when the sequence of DNA is known
Konika Naval
August 31, 2025Degenerate primers are used even when the sequence of DNA is known
Kanica Sunwalka
August 31, 2025Degenerated primers are random primers used even when dna seq is known
Meenakshi Choudhary
September 3, 2025Degenerate primers are used even when the sequence of DNA is known.
Muskan Yadav
September 8, 2025Degenerate primers are used even when the sequence of DNA is known is the correct answer