7 public questions tagged with this topic.
DNA probes are used in molecular diagnostics to detect specific DNA sequences by hybridization.
Ref: NCERT Class 12 Biology Chapter 10: Biotechnology and its Applications Biosafety and Overview of Applications
Autoradiography is used to detect specific DNA sequences by hybridizing them with radioactive probes.
Ref: NCERT Class 12 Biology Chapter 10: Biotechnology and its Applications Medical Applications - Molecular Diagnosis - PCR ELISA
TaqMan chemistry exploits the 5' to 3' exonuclease activity of Taq polymerase. The linear dual-labeled probe hybridizes to template during annealing, and during extension polymerase cleaves the probe, separating fluorophore from quencher and producing irreversible fluorescence accumulation. Molecular beacons instead rely on reversible hybridization without cleavage; their stem-loop opens at annealing when bound to target and closes after denaturation, yielding transient fluorescence at annealing phase. TaqMan does not possess stem-loop structure, nor does it bind minor grooves. The key mechani
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.
Western blotting is inherently unsuitable for DNA detection because it employs antibodies raised against polypeptide epitopes and conditions optimized for protein immobilization and immunorecognition. DNA molecules lack immunoglobulin recognition sites and are not detected by anti-protein antibodies. Furthermore, no nucleic acid probe hybridization step is incorporated. Compatible DNA analysis methods include native or denaturing polyacrylamide gel electrophoresis stained with ethidium bromide, agarose electrophoresis, denaturing high-performance liquid chromatography separating based on melti
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.
Southern blotting is classical technique established for specific DNA detection within heterogeneous genomic DNA population. Purified DNA is cut with restriction endonucleases, size-fractionated on agarose, denatured to single-stranded form, transferred to membrane via capillary action, and hybridized with labeled probe complementary to gene of interest. Visualization confirms presence, copy number, size, and rearrangement status of locus. Applications include mutation detection, RFLP mapping, and transgene integration analysis. In contrast, Northern blotting detects mRNA, Western detects prot
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.
Southern blotting, invented by Edwin Southern in 1975, remains the definitive method for detection of specific DNA sequences. Genomic DNA is fragmented with restriction endonucleases, resolved by agarose gel electrophoresis according to size, denatured under alkaline conditions to single strands, and capillary transferred to nitrocellulose or nylon membrane. Immobilized fragments are hybridized with radioactively or non-radioactively labeled complementary probes. Detection reveals fragment length polymorphism, enabling genotyping, DNA fingerprinting, transgene confirmation, and analysis of gen
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.
Detection of radioactively labeled DNA fragments relies on emission of beta particles from isotopes such as 32P incorporated at termini. Gel electrophoresis separates fragments by size but does not detect radioactivity itself. Spectrometry and colorimetry measure absorbance or color development for quantification, not spatial band position. Autoradiography places dried gel against photographic film; radioactive decay exposes silver halide crystals producing dark bands corresponding precisely to labeled fragment migration positions. This method provides high sensitivity for low abundance labele
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.
