Rhodamine derivatives are not sufficiently stable to survive conventional deprotection and these must be attached to amino-modified oligonucleotides using post-synthesis labelling techniques. Because Tetramethyl Rhodamine (TAMRA) is not base stable, the procedure to cleave and deprotect the labelled oligonucleotide must be carefully considered. Using the UltraMILD monomers and deprotection with potassium carbonate in methanol, TAMRA oligonucleotides can be fairly conveniently isolated. To streamline the preparation of TAMRA oligos, we offer 3'-TAMRA CPG for 3' labelling and TAMRA-dT for labelling within the sequence. We also offer TAMRA NHS ester for labelling amino-modified oligonucleotides.
Coupling: Dissolve this product in anhydrous acetonitrile:THF (9:1). Allow 15 minutes for the product to go into solution. A 6 minute coupling time is recommended. Use either dmf-dG (10-1029-xx) with standard monomers and Cap A or use UltraMILD monomers (dA: 10-1601-xx, dC: 10-1015-xx, dG: 10-1621-xx, dT: 10-1030-xx ) and UltraMILD Cap Mix A (40-4210-xx/ 40-4212-xx)
Deprotection: If standard monomers were used, deprotect in tert-butylamine/water 1:3 (v/v) for 6 hours at 60 °C. If UltraMILD monomers were used, deprotect in 0.05M Potassium Carbonate in Methanol, 4 hours at Room Temperature. See Technical Bulletin for details (http://www.glenresearch.com/Technical/TB_UltraMild_Deprotection.pdf).