The DNA tethering kit is a highly efficient system for tethering a DNA sequence of interest (SOI) in optical tweezers and studying the dynamics of DNA-binding proteins. Using the kit, the SOI can be ligated to two DNA handles. Each DNA handle is 6.3 kb long (exact size: 6,298 bp; download the sequence here), modified with 3 biotin moieties on one end and a 4-nucleotide single-stranded DNA overhang on the other end. To enable ligation of the SOI to the DNA handles, the SOI requires overhangs complementary to those of the DNA handles. The overhangs can be introduced into the SOI by digestion with a TypeIIS restriction enzyme, following the guidelines described in the online protocol.
Assembly of multiple DNA fragments using Golden Gate Assembly is also compatible with the DNA tethering kit. DNA fragments with a combined length of up to 24 kb have been ligated to 6.3 kb handles. For questions on using Golden Gate Assembly with the DNA tethering kit, please reach out to store@lumicks.com.
The ligation is designed so that DNA handle 1 will be ligated at the 5′ end of the sequence of interest, and DNA handle 2 will be ligated at the 3′ end. Additionally, the kit offers different versions of handles with or without ATTO fluorophores. The options for handle 1 include with ATTO 647N or without a fluorophore, while for handle 2, the options are with ATTO 647N, ATTO 488, or without a fluorophore. The combination of two handles without fluorophores is offered for a discounted early release price via this product page.
In addition to the DNA handles, the kit includes a DNA Ligase enzyme, 10x DNA Ligase buffer, a 3 kbp control DNA with overhangs (exact size: 3,065 bp), and a plasmid (discover the pUC-LUMICKS plasmid here).
The kit enables 4 DNA Ligase reactions. Each DNA ligase reaction can be used for at least 5 experimental sessions on the C-Trap. Please note that the current price of the product is an temporary, introduction offer.
Estimated 1-4 weeks delivery time.
