Nucleic acid amplification kits for specific detection of e. coli o157:h7 without co-detection of e. coli o55:h7

Abstract

Disclosed are assays, methods and kits for the specific detection of E. coli O157:H7 and not E. coli O55:H7 from complex food matrices, water, a beverage sample, a fermentation broth, a forensic sample, an environmental sample (e.g., soil, dirt, garbage, sewage, air, or water), including food processing and manufacturing surfaces, or a biological sample.

1 - 67 . (canceled) 
     
     
         68 . A kit for the detection of  E. coli  O157:H7 without the co-detection of  E. coli  O55:H7 in a sample comprising:
 a. at least one of a first pair of PCR primers selected from: SEQ ID NO:1 and SEQ ID NO: 2, SEQ ID NO:1 and SEQ ID NO:4, SEQ ID NO:1 and SEQ ID NO:5, and SEQ ID NO:1 and SEQ ID NO:6, and full complements thereof;   b. at least one of a second pair of PCR primers selected from SEQ ID NO:7-8, SEQ ID NO:10-11, SEQ ID NO:13-14, SEQ ID NO:16-17, SEQ ID NO:19-20, SEQ ID NO:22-23, SEQ ID NO:25-26, SEQ ID NO:28-29, SEQ ID NO:31-32, SEQ ID NO:34-35, SEQ ID NO:37-38, SEQ ID NO:40-41, SEQ ID NO:43-44, SEQ ID NO:46-47, SEQ ID NO:49-50, SEQ ID NO:52-53, SEQ ID NO:55-56, SEQ ID NO:59 and SEQ ID NO:56, SEQ ID NO:61-62, SEQ ID NO:64-65, SEQ ID NO:67-68, SEQ ID NO:70-71, SEQ ID NO:73-74, SEQ ID NO:76-77, SEQ ID NO:79-80, SEQ ID NO:82-83, SEQ ID NO:85-86, and SEQ ID NO:88-89 and full complements thereof; and   c. a polymerase.   
     
     
         69 . The kit of  claim 75  wherein said first probe further comprises a first label and said second probe further comprises a second label. 
     
     
         70 . The kit of  claim 69 , wherein both labels are selected from a dye, a radioactive isotope, a chemiluminescent label, and an enzyme. 
     
     
         71 . (canceled) 
     
     
         72 . The kit of  claim 70 , wherein the dye comprises a fluorescein dye, a rhodamine dye, or a cyanine dye. 
     
     
         73 . The kit of  claim 71 , wherein the dye is a fluorescein dye. 
     
     
         74 . The kit of  claim 69 , wherein said first probe is labeled with FAM™ dye and said second probe is labeled with VIC® dye. 
     
     
         75 . The kit of  claim 68 , further comprising:
 a first probe of SEQ ID NO:3 or a full complements thereof for use with selected said first pair of PCR primers and a second probe for use with selected said second pair of PCR primers selected from SEQ ID NO:9, SEQ ID NO:12, SEQ ID NO:15, SEQ ID NO:18, SEQ ID NO:21, SEQ ID NO:24, SEQ ID NO:27 SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:36, SEQ ID NO:39, SEQ ID NO:42, SEQ ID NO:45, SEQ ID NO:48, SEQ ID NO:51, SEQ ID NO:54, SEQ ID NO:57, SEQ ID NO:58, SEQ ID NO:60, SEQ ID NO:63, SEQ ID NO:66, SEQ ID NO:69, SEQ ID NO:72, SEQ ID NO:75, SEQ ID NO:78, SEQ ID NO:81, SEQ ID NO:84, SEQ ID NO:87, and SEQ ID NO:90 and full complements thereof.   
     
     
         76 . The kit of  claim 68 , wherein the polymerase is a DNA polymerase. 
     
     
         77 . The kit of  claim 68 , further comprising one or more components selected from, one or more buffers, a nucleotide triphosphate, an intercalating dye and instructions for using the kit.