Enzymatic process

Enzymatic process

  • CN 104,126,018 B
  • Filed: 12/28/2012
  • Issued: 09/14/2021
  • Est. Priority Date: 12/29/2011
  • Status: Active Grant
First Claim
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1. A method of characterizing a target polynucleotide, comprising:

  • (a) contacting a target polynucleotide with a transmembrane pore and a RecD helicase such that the target polynucleotide moves through the pore and the RecD helicase controls movement of the target polynucleotide through the pore; and

    (b) obtaining one or more measurements as the polynucleotide moves relative to the pore, wherein the measurements are representative of one or more characteristics of the target polynucleotide and thereby characterise the target polynucleotide,wherein the process is carried out using a salt concentration of 0.3M to 0.4M;

    wherein the transmembrane pore is selected from the group consisting of hemolysin, leukocidin, Mycobacterium smegmatis porin A (MspA), outer membrane porin F (OmpF), outer membrane porin G (OmpG), outer membrane phospholipase A, Neisseria autotransporter lipoprotein (NalP) and WZA; and

    isWherein the RecD helicase comprises the amino acid modular sequence X1-X2-X3-X4-X5- (X6)3-Q-X7(SEQ ID NO;

    16) wherein X1 is Y, W or F, X2 is A, T, S, M, C or V, X3 is any amino acid, X4 is T, N or S, X5 is A, T, G, S, V or IX6 is any amino acid and X7 is G or S.

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