Method for the detection of diagnostic rna

Abstract

Methods for the direct detection of diagnostic target RNA have been developed, which obviate the need for time consuming RNA purification and isolation procedures.

1 - 22 . (canceled) 
     
     
         21 . A kit for the direct detection of a diagnostic target RNA from a bacteria, the kit comprising:
 A) instructions describing a method for the direct detection of a diagnostic target RNA from a bacteria comprising the steps of:
 a) providing a sample comprising at least a portion of at least one bacterial cell, said portion of said cell comprising a diagnostic target RNA; 
 b) contacting the sample with an RT-PCR composition to form an RT-PCR reaction mixture; 
 c) thermocycling the RT-PCR reaction mixture, whereby a diagnostic target DNA product is amplified; and 
 d) detecting the amplified diagnostic target DNA product of (c), with the proviso that no step includes RNA purification and wherein the sample of (a) does not contain an exogenously added RNAase inhibitor; 
 and 
   B) an RT-PCR composition, wherein the RT-PCR composition comprises at least one of the following elements: at least one primer complementary to the diagnostic target RNA; a heat activated thermostable polymerase; a reverse transcriptase enzyme; dNTPs; or appropriate buffers.   
     
     
         22 . A kit for the direct detection of a diagnostic target RNA from a bacteria, the kit comprising:
 A) instructions describing a method for the direct detection of a diagnostic target RNA from a bacteria, comprising the steps of:
 a) providing a sample comprising at least a portion of at least one bacterial cell, said portion of said cell comprising a diagnostic target RNA; 
 b) contacting the sample with an RT reaction composition to form an RT reaction mixture under conditions, whereby a diagnostic DNA product is transcribed from the RNA; 
 c) detecting the diagnostic target DNA product, with the proviso that no step includes RNA purification and wherein the sample of (a) does not contain an exogenously added RNAase inhibitor; and 
   B) an RT reaction composition, wherein the RT composition comprises at least one of the following elements: at least one nucleic acid primer complementary to the diagnostic target RNA; a reverse transcriptase enzyme; dNTPs; appropriate buffers; detection dyes; or detection probes.   
     
     
         23 . A kit according to  claim 22 , wherein the method further comprises after step (b), the steps of:
 contacting the diagnostic DNA product with a PCR composition to result in a PCR reaction mixture; and   thermocycling the resultant PCR reaction mixture, whereby the diagnostic target DNA product is amplified.   
     
     
         24 . A kit according to  claim 22 , wherein the detecting step of the method comprises incorporating or attaching a label to the diagnostic target DNA product.