Method, lysis solution and kit for selectively depleting animal nucleic acids in a sample

Abstract

The present invention relates to a method for selectively depleting animal nucleic acids from non-animal nucleic acids in a sample, which comprises animal cells and at least one further type of cells, selected from microbial cells and plant cells or a combination thereof, to a lysis solution A to be used in and to a kit to carry out said method as well as to the use of a particular silica membrane as both, a filtration matrix for separating essentially intact microbial and/or plant cells from lysed animal cells and an adsorption matrix for nucleic acids, in particular in a method according to the present invention.

1 . A lysis solution, comprising at least one saponin, a water-soluble salt selected from the group consisting of acetates, sulfates, glutamates, and any mixture thereof; a viscosity modifying agent selected from the group consisting of sucrose, glucose, and a mixture thereof; and optionally a polyanionic sulfonate. 
     
     
         2 . The lysis solution of  claim 1 , wherein the at least one saponin is present in an amount ranging from about 0.01 to about 15% (w/v), based on the whole lysis solution, wherein the aforementioned concentration range refers to the total amount of saponin. 
     
     
         3 . The lysis solution of  claim 1 , wherein the water-soluble salt is present in a concentration ranging from 10 to 1.000 mM, wherein the aforementioned concentration range refers to the total amount of the water-soluble salt based on the whole lysis solution. 
     
     
         4 . The lysis solution of  claim 1 , wherein the viscosity modifying agent is present in a concentration ranging from 10 to 1.000 mM, wherein the aforementioned concentration range refers to the total amount of the viscosity modifying agent based on the whole lysis solution. 
     
     
         5 . The lysis solution of  claim 1 , wherein the polyanionic sulfonate is present in a concentration ranging from 0.001 to 50 mM, wherein the aforementioned concentration range refers to the total amount of the polyanionic surfactant based on the whole lysis solution. 
     
     
         6 . The lysis solution of  claim 1 , wherein said lysis solution is chaotrope-free and further comprises at least one additional nonionic surfactant in an amount from 0.01 to 15% (w/v) based on the whole lysis solution. 
     
     
         7 . The lysis solution of  claim 6 , wherein the additional nonionic surfactant is selected from the group consisting of a sorbitan ester of fatty acids, a polyoxyethylene sorbitan ester of fatty acids, a polyoxyalkylene ether of fatty alcohols, a polyoxyalkylene ether of alkylphenols, a poloxamer, and mixtures of any thereof. 
     
     
         8 . A kit for selectively depleting animal nucleic acids from a sample comprising animal cells and microbial and/or plant cells, the kit comprising:
 (i) the lysis solution of  claim 1 , and   (ii) optionally, an endonuclease capable of degrading both RNA as well as DNA or a mixture of at least one DNase and at least one RNase, and   (iii) optionally, a filter for separating intact microbial and/or plant cells from a liquid part of the sample, wherein the liquid part of the sample includes digested animal nucleic acids, by retaining the microbial and/or plant cells while the liquid part of the sample passes through the filter as a filtrate.   
     
     
         9 . The kit according to  claim 8 , further comprising at least one of the following components: instructions for using the kit, chromatographic means for purifying, concentrating and/or isolating microbial and/or plant cells or nucleic acids, agents or buffers for lysing the microbial and/or plant cells, binding mediators or binding buffers, washing buffers, elution buffers, further means for filtering and/or centrifuging the sample, or any combination thereof. 
     
     
         10 . The kit according to  claim 8 , wherein the filter of (iii) fulfills at least one of the following conditions:
 a) the filter comprises at least one part having a pore size being in the range of 0.1-1.5 μm, preferably 0.2-1.0 μm, more preferred 0.3-0.8 μm (particularly 0.25+/−0.05−0.8 μm) and most preferred 0.5-0.7 μm; and   b) the filter comprises polyester, polyethersulfone (PES), cellulose, cellulose acetate (CA), mixed cellulose esters (MCE), polytetrafluoroethylene (PTFE), polyamide (PA), Nylon, polypropylene (PP), ceramics, glass fibers, silica, or any combination or mixture thereof.