UNIT 8.8 Single-Nucleotide Sequence Discrimination In Situ Using Padlock Probes
Published Online: 1 MAY 2001
Copyright © 2003 by John Wiley and Sons, Inc.
Lab Protocol Title
Current Protocols in Cytometry
How to Cite
Nilsson, M., Landegren, U. and Antson, D.-O. 2001. Single-Nucleotide Sequence Discrimination In Situ Using Padlock Probes. Current Protocols in Cytometry. 16:8.8:8.8.1–8.8.12.
- Published Online: 1 MAY 2001
- Published Print: APR 2001
Standard fluorescence in situ hybridization (FISH) techniques using cloned probes are limited in their ability to distinguish between closely similar DNA sequences because long hybridization probes are not detectably destabilized by single mismatched base pairs. This problem has been addressed by using short allele-specific oligonucleotide probes whose hybridization to target sequences is more sensitive to mismatches. This revised and expanded unit presents protocols for discrimination between closely similar DNA sequences in situ. The discussion of probe synthesis has been greatly expanded and an Alternate Protocol 1 added for enzymatic probe ligation at low probe concentration. A new Support Protocol describes enzymatic probe synthesis.