NH3/MeOH 30 min at r.t., then 1ml of 7M NH3/MeOH, 5h at 65oC, followed by TOM deprotection** 0.4ml of 4M NH3/MeOH 30 min at r.t., then 1ml of 2M Me-NH2/ MeOH, 30 min at 65oC, followed by TOM deprotection** 0.4 ml 32% NH3/H2O for 5h at 65oC, followed by TOM deprotection** 0.4ml of 32% NH3/H2O 30 min at r.t., then 1ml of 40% Me-NH2/ H2O, 30 min at 65oC, followed by TOM deprotection** 0.4ml of 4M NH3/MeOH 30 min at r.t., then 1ml of 7M NH3/MeOH, 5h at 65oC, followed by TOM deprotection** 0.4ml of 4M NH3/MeOH 30 min at r.t., then 1ml of 2M Me-NH2/ MeOH, 30 min at 65oC, followed by TOM deprotection** 0.4 ml 32% NH3/H2O for 5h at 65oC, followed by TOM deprotection** 0.4ml of 32% NH3/H2O 30 min at r.t., then 1ml of 40% Me-NH2/ H2O, 30 min at 65oC, followed by TOM deprotection**
# Two strands of siRNA were synthesized in this study. The sense strand 5′-AGUCGCCUCGAAGAUACACtt-3′ was synthesized on CPG-based support 5a and 6, and the antisense strand 5′-GUGUAUCUUCGAGGCGACUtt-3′ was synthesized on polymeric supports 4 and 5b. Uppercase letters are given for the Ribonucleoside units and the lowercase t for the thymidine unit. ## The yield of target oligonucleotide, generated from the CPG Universal Support 5a with NH3/MeOH, was taken as 100% for oligos 1-4. The yield of target oligonucleotide, generated from the polystyrene Universal Support 5b with NH3/MeOH, was taken as 100% for the oligos 5-8. Contents of oligonucleotides in the crude mixtures were determined by ion-exchange HPLC. * The structure of Universal Support 6 was not disclosed by the manufacturer. ** 0.5 ml of DMSO + 0.16 ml of HF/TEA, 65 , 30 min.; cool on ice and quench with 1 ml 0.1M Na acetate (sterile!!!), pH 5.2, 65 , 30 min.; cooled on ice, desalted and analyzed.
Seq 2 was synthesized on 1000 dG-CPG and Seq 1,3,4-6 on 1000 Universal Support 5a. *RP HPLC was used for oligonucleotide purification. ** Ion-exchange HPLC, followed by RP HPLC were used for oligonucleotide purification. 7
SYNTHESIS OF LONGER DNA OLIGOMERS ON UNIVERSAL SUPPORTS While universal supports 5a and 5b performed best for DNA and RNA synthesis, is it reasonable to expect a universal support to be compatible with the synthesis of longer DNA oligos An experiment was set up to compare the yield and purity of oligos prepared on a 1000CPG version of Support 5a with the synthesis on a conventional 1000deoxynucleoside support.86445-22-9 web The results of this experiment are collected in Table 4. The product oligonucleotides Seq 1, 2, 4, 5 were purified DMT-on by reverse phase HPLC.1414811-15-6 manufacturer The other two oligos, Seq 3, 6, were first purified DMT-off by ion-exchange HPLC followed by reverse phase HPLC.PMID:30846224 Yields were determined by measuring AU260. As detailed in Table 4, it is clear that the universal support 5a is compatible with the synthesis of long oligos and good quality products can be obtained in good yield.
CONCLUSION The main impediment to the universal adoption of a universal support has been the aggressively basic conditions required to complete the elimination reaction to release the terminal hydroxyl group. The standard reagents used in oligonucleotide deprotection are ammonium hydroxide and aqueous methylamine, which are popular since they are completely volatile. Using these reagents to carry out the elimination reaction requires either high temperature, with attendant high pressure, or extended reaction times. In addition, lithium chloride has been used to speed up the elimination reaction. However, the addition of salts to the dep.MedChemExpress (MCE) offers a wide range of high-quality research chemicals and biochemicals (novel life-science reagents, reference compounds and natural compounds) for scientific use. We have professionally experienced and friendly staff to meet your needs. We are a competent and trustworthy partner for your research and scientific projects.Related websites: https://www.medchemexpress.com
