Cdpcho

Restriction enzymes and DNA polymerases were obtained from New England Biolabs (Ipswich, MA, USA). Isopropyl β-D-1-thiogalactopyranoside (IPTG) was obtained from Fisher Scientific GmbH (Schwerte, Germany). Nickel-nitrilotriacetic acid (Ni-NTA) was from Qiagen (Düsseldorf, Germany), protease inhibitor cocktail tablets were purchased from Roche (Basel, Switzerland). CTP, CDPCho, Sypro Orange, inorganic pyrophosphatase, purine nucleoside phosphorylase, DNA purification kit and antibiotics were purchased from Sigma-Aldrich (St Louis, MO, USA). Phosphocholine chloride sodium salt hydrate (further termed as ChoP) was from TCI Europe N.V. (Antwerp, Belgium). MESG (7-methyl-6-thioguanosine) was obtained from Berry and Associates (Dexter, MI, USA). All other chemicals were of analytical grade of the highest purity available.

CTP was purchased from Thermo Fisher Scientific (Mississauga, ON, Canada). Chelex 100 sodium form (50–100 mesh), AEP, P-Cho, and CDP-Cho were purchased from Sigma-Aldrich (Oakville, ON, Canada). P-Cho was treated with Chelex 100 resin as previously described prior to use in order to remove calcium, which inhibits the enzyme^{32 (link)}. All other chemicals were used without further purification. DNA sequencing was performed at The Centre for Applied Genomics at the Hospital for Sick Children (Toronto, ON, Canada). HR–ESI–MS was performed at the Alberta Glycomics Centre (Edmonton, AB, Canada). HPLC analysis was performed using a Prominence LC-20AT system equipped with a diode array detector (Shimadzu, Kyoto, Japan). FPLC purifications were performed using an NGC™ Chromatography System (Bio-Rad, Mississauga, ON). NMR spectra were recorded on an Agilent DD2 operating at 400 MHz for ¹H and 100 MHz for ¹³C. A Varian Unity Inova operating at 121 MHz was used to record ³¹P NMR spectra.