Glen Report 22.29: Technical Brief - Crosslinking with Click Chemistry

The click process is rapidly becoming the method of choice for key organic reactions useful in oligonucleotide modification.1 The advent of Copper (Cu1) catalyzed [3+2] azide-alkyne click chemistry (CuACC) has been the main reason for the surge in interest in this chemistry. CuACC reactions have simply improved the speed and performance of click chemistry. At the same time, the specificity of the reaction between an alkyne and an azide means that side reactions are very unlikely even in complex biological systems.

FIg 1
Figure 1: Crosslinking with click chemistry

One of our most common questions from researchers is how to create a specific crosslink between the strands of an oligonucleotide duplex. Most procedures use light to create a crosslink using a free radical mechanism. Photolytic crosslinking is typically low yielding with many side reactions possible. Figure 1 shows click chemistry being used2 to form such an internucleotide crosslink. One DNA strand is simply modified using C8-Alkyne-dT (10-1540). The complementary strand is first modified using Amino-Modifier C6-dT (10-1039). A post-synthesis conjugation with Azidobutyrate NHS ester (50-1904) leads to the azido-modified strand. A subsequent CuACC click reaction leads to the intranucleotide crosslink in high yield.

In a recent article, this crosslinking technique was extended to hairpin and hammerhead ribozymes.3


  1. A.H. El-Sagheer, and T. Brown, Chem Soc Rev, 2010, 39, 1388-405.
  2. P. Kocalka, A.H. El-Sagheer, and T. Brown, Chembiochem, 2008, 9, 1280-1285.
  3. A.H. El-Sagheer, and T. Brown, Proc Natl Acad Sci U S A, 2010, 107, 15329-34.

Product Information

Click and Copper-free Click Chemistry