Agilent Technologies has been granted a patent for mutant polymerases that have an improved ability to incorporate modified nucleotides, including 3′-OH unblocked reversible terminators. These polymerases can be used in various applications such as polynucleotide sequencing, primer extension reactions, and enzymatic oligonucleotide synthesis. The patent also includes a composition comprising a 3′-OH unblocked reversible terminator and a mutant polymerase with specific amino acid mutations. GlobalData’s report on Agilent Technologies gives a 360-degree view of the company including its patenting strategy. Buy the report here.
According to GlobalData’s company profile on Agilent Technologies, Microfluidics automation was a key innovation area identified from patents. Agilent Technologies's grant share as of September 2023 was 52%. Grant share is based on the ratio of number of grants to total number of patents.
Improved mutant polymerases for incorporating modified nucleotides
A recently granted patent (Publication Number: US11773380B1) discloses a composition and methods for nucleotide sequencing. The composition includes a 3'-OH unblocked reversible terminator and a mutant polymerase. The mutant polymerase has specific amino acid mutations at positions functionally equivalent to amino acid positions at K477, A486, and Y546 in Pfu polymerase. The mutant polymerase can also have additional mutations at other positions, such as F494. The mutant polymerase can be derived from Pyrococcus or Thermococcus polymerase.
The patent also describes methods for incorporating a nucleotide to a priming strand and polynucleotide sequencing. In the method of incorporating a nucleotide, the priming strand is contacted with a nucleotide and the mutant polymerase under conditions suitable for an incorporation reaction. The nucleotide used can be a 3'-OH unblocked reversible terminator. The method of polynucleotide sequencing involves forming a duplex with a template and a priming strand, combining the priming strand with a reversible terminator nucleotide and the mutant polymerase, incorporating the reversible terminator at the 3'-end of the priming strand in a template-dependent reaction, and identifying the incorporated reversible terminator nucleotide to determine the sequence of the template. This method can be repeated multiple times for accurate sequencing.
Another composition disclosed in the patent includes a priming strand, a 3'-OH-unmodified reversible terminator, and a mutant polymerase. The mutant polymerase has specific amino acid mutations at positions functionally equivalent to amino acid positions at Y546, L409, and A486 in Pfu polymerase. The mutant polymerase can also have additional mutations at other positions, such as L270, E330, Q332, L333, P451, L453, L457, E476, L489, L490, N492, F494, Y497, and E581. The mutant polymerase can be derived from Pyrococcus or Thermococcus polymerase and has an incorporation activity at least 2-fold higher than a DNA polymerase of SEQ ID NO:11.
Overall, this patent provides compositions and methods for nucleotide sequencing using specific mutant polymerases and reversible terminators. These innovations can potentially improve the accuracy and efficiency of nucleotide sequencing techniques.