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#restriction enzyme

18 public questions tagged with this topic.

ZOOLOGY

Which enzyme is used to produce sticky ends in a DNA fragment?

Restriction endonucleases cut DNA at specific sequences, generating sticky ends for recombination. This follows from latest NCERT 2026-27 principle explaining the concept clearly for NEET students in simple steps as per rationalized syllabus.

Ref: NCERT Biology Textbook - Latest Edition for Academic Session 2026-27 (Zoology section, Rationalized Textbook for Class XI and XII), Chapter: Biology - Zoology portion (Latest NCERT Textbooks for Academic Session 2026-27 - Rationalized Edition for Class XI and XII), Topic: Structural organization, physiology, human health and related concepts as per latest syllabus.

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ZOOLOGY

Which enzyme is used to cut DNA at specific sites during genetic manipulation?

Endonuclease enzymes cut DNA at specific recognition sites during genetic manipulation. This follows from NCERT principle where the relation explains the outcome clearly for students in simple steps.

Ref: NCERT Biology Textbook for Class XI and XII (Zoology section), Chapter: Principles of Inheritance, Molecular Basis of Inheritance and Biotechnology, Topic: Genetics, DNA techniques and applications.

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RDT Basics

Restriction enzyme SalI cuts at:

SalI is a Type II restriction endonuclease isolated from Streptomyces albus that recognizes palindromic hexamer 5'-G^TCGAC-3' and cleaves between G and TCGAC to generate four-nucleotide 5' overhang with sequence TCGA. The staggered cut produces cohesive termini that anneal efficiently, enabling ligation with other fragments sharing TCGA overhangs, notably those produced by XhoI. Cleavage requires magnesium as cofactor and is inhibited if the recognition site overlaps Dam-methylated GATC leading to methylation-dependent blockage. SalI remains widely used for mapping large plasmids, constructing genomic libraries, and directional subcloning applications.

Ref: NCERT Biology Class XII Principles on Klenow fill-in labeling, Lehninger Chapter 9 DNA cloning techniques, and Molecular Cloning by Sambrook Chapter 10 documenting end-labeling of cohesive termini.

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RDT Basics

Enzyme that produces sticky ends at C/TCGAG:

XhoI, derived from Xanthomonas holcicola, is a Type II restriction enzyme recognizing hexameric palindrome 5'-C^TCGAG-3' and cutting after first base to leave four-base 5' overhang TCGA. Cleavage requires magnesium and produces cohesive ends compatible with SalI, compatible with XhoI-generated fragments, and useful for creating complementary junctions. The enzyme cuts rarely in AT-rich genomes due to GC-rich recognition, making it attractive for constructing expression cassettes, removing promoters, and performing directional cloning where unique sites are needed to maintain reading frame and transcription orientation.

Ref: NCERT Biology Class XII Principles on Klenow fill-in labeling, Lehninger Chapter 9 DNA cloning techniques, and Molecular Cloning by Sambrook Chapter 10 documenting end-labeling of cohesive termini.

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RDT Basics

Enzyme producing blunt ends at CCC/GGG:

SmaI, purified from Serratia marcescens, recognizes palindromic sequence 5'-CCC^GGG-3' and hydrolyzes phosphodiester bond between third cytosine and first guanine, denoted CCC^GGG, generating blunt-ended fragments lacking single-stranded extensions. Blunt-end ligation demands higher concentrations of T4 DNA ligase and conditions stabilizing ends, contrasting with efficient sticky-end ligation. SmaI sites are GC-rich and often located in CpG islands, and methylation of internal CpG at C5 can inhibit cleavage due to overlap with eukaryotic methylation patterns, influencing mapping and subcloning strategies in genomes and vectors.

Ref: NCERT Biology Class XII Principles on Klenow fill-in labeling, Lehninger Chapter 9 DNA cloning techniques, and Molecular Cloning by Sambrook Chapter 10 documenting end-labeling of cohesive termini.

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