Hi Nurul,

Personally, I have done all my cloning so far with desalted primers and have never had any issues. I think you can save some money here by going for the cheaper option.

Good Luck!

-Jan

P.S: I always add 6 bases to the restriction site for efficient digestion.

Dear Nurul Nadirah Razali

Regarding the number of bases to be added before the restriction site it may chenge on the basis of the specific enzime. You can find sequences suggested by NEB in the following link:

https://international.neb.com/-/media/nebus/files/chart-image/cleavage_olignucleotides_old.pdf?rev=c2f94e1cdcd549c5bf8fdb59f7b63f67&hash=0C83C8F3C59132BE6A5B9A6D4050E3A0

As you can see, form the file it seems that for both Xhoi and Xba 3 bases are anough to reach the best activity

if you are interested you can also find some example of primer desing on the folllowing link:

https://www.blogger.com/video.g?token=AD6v5dzxz85z69as_qBJCTiisMtVv4q-Q0ChLHJHqSwNRA_FWtuB9p36qi4nQxd4b3OB2cxClbV6BmNDgkIdwfDUZqcDdOrGBSsjcuGTQKHAIEJPIrzIbIxwHrRRfZQf_UG0xZwwe2I

on my blog (ProteoCool)

good luck

Manuele

I agree that HPLC purification is not needed for successfull cloning. Standard quality primers without any additional purification work very well. Some companies may tell you that they "strongly recommend additional purification for cloning". I found this to be untrue. HPLC purification adds significantly to price and time of manufacturing. So, getting rid of this (not needed) step would be imo the correct decision.

in my opinion HPLC purification not required, I did so many times cloning with without HPLC purified primers