摘要 |
A method for amplification and capture of nucleic acid sequences can include the steps of annealing a forward primer to a DNA or RNA template in a first reaction vessel that includes fewer than four different dNTPs; extending the forward primer with the dNTPs to form an extended primer that terminates when an omitted dNTP is required for further extension of the forward primer; releasing the extended primer; exponentially amplifying the extended primer in a second reaction vessel that includes a reverse primer, four different dNTPs and a capture probe, the capture probe including n oligonucleotides, wherein fewer than n of the oligonucleotides are locking nucleic acids; and concurrently capturing one of the extended primers in the second reaction vessel with the capture probe while amplifying the extended primer. Further, in certain embodiments, the steps of annealing, extending and releasing occur at a first reaction temperature that is substantially isothermal and in the absence of a helicase. In addition, or alternatively, the steps of exponentially amplifying and capturing occur at a second reaction temperature that is substantially isothermal.
|