Pcr primer set detecting severe acute respiratory syndrome (sars)-coronavirus, method and kit for detecting sars-coronavirus using the same

Abstract

Provided are SARS-CoV detection PCR primers having nucleotide sequences as set forth in SEQ ID NOS: 1-46, a method for detecting SARS-CoV using the primers, and a SARS-CoV detection kit including the primers.

1 . A PCR primer set selected from the group consisting of the following primer sets:
 (a) a primer set (GS-SARS02) comprising a primer consisting of SEQ ID NO: 3 and a primer consisting of SEQ ID NO: 4;   (b) a primer set (GS-SARS03) comprising a primer consisting of SEQ ID NO: 5 and a primer consisting of SEQ ID NO: 6;   (b) a primer set (GS-SARS04) comprising a primer consisting of SEQ ID NO: 7 and a primer consisting of SEQ ID NO: 8;   (c) a primer set (GS-SARS05) comprising a primer consisting of SEQ ID NO: 9 and a primer consisting of SEQ ID NO: 10;   (d) a primer set (GS-SARS06) comprising a primer consisting of SEQ ID NO: 11 and a primer consisting of SEQ ID NO: 12;   (e) a primer set (GS-SARS07) comprising a primer consisting of SEQ ID NO: 13 and a primer consisting of SEQ ID NO: 14;   (f) a primer set (GS-SARS08) comprising a primer consisting of SEQ ID NO: 15 and a primer consisting of SEQ ID NO: 16;   (g) a primer set (GS-SARS09) comprising a primer consisting of SEQ ID NO: 17 and a primer consisting of SEQ ID NO: 18;   (h) a primer set (GS-SARS10) comprising a primer consisting of SEQ ID NO: 19 and a primer consisting of SEQ ID NO: 20;   (i) a primer set (GS-SARS11) comprising a primer consisting of SEQ ID NO: 21 and a primer consisting of SEQ ID NO: 22;   (j) a primer set (GS-SARS12) comprising a primer consisting of SEQ ID NO: 23 and a primer consisting of SEQ ID NO: 24;   (k) a primer set (GS-SARS13) comprising a primer consisting of a SEQ ID NO: and a primer consisting of SEQ ID NO: 26;   (l) a primer set (GS-SARS14) comprising a primer consisting of as set forth in SEQ ID NO: 27 and a primer consisting of as set forth in SEQ ID NO: 28;   (m) a primer set (GS-SARS15) comprising a primer consisting of SEQ ID NO: 29 and a primer consisting of SEQ ID NO: 30;   (n) a primer set (GS-SARS16) comprising a primer consisting of SEQ ID NO: 31 and a primer consisting of SEQ ID NO: 32;   (o) a primer set (GS-SARS17) comprising a primer consisting of SEQ ID NO: 33 and a primer consisting of SEQ ID NO: 34;   (q) a primer set (GS-SARS18) comprising a primer consisting of SEQ ID NO: 35 and a primer consisting of SEQ ID NO: 36;   (r) a primer set (GS-SARS19) comprising a primer consisting of SEQ ID NO: 37 and a primer consisting of SEQ ID NO: 38;   (s) a primer set (GS-SARS20) comprising a primer consisting of SEQ ID NO: 39 and a primer consisting of SEQ ID NO: 40;   (t) a primer set (GS-SARS21) comprising a primer consisting of SEQ ID NO: 41 and a primer consisting of SEQ ID NO: 42;   (u) a primer set (GS-SARS22) comprising a primer consisting of SEQ ID NO: 43 and a primer consisting of SEQ ID NO: 44; and   (v) a primer set (GS-SARS23) comprising a primer consisting of SEQ ID NO: 45 and a primer consisting of SEQ ID NO: 46,   wherein the primer set specifically amplifies a target region of Severe Acute Respiratory syndrome corona virus (SARS-CoV) in a polymerase chain reaction (PCR) and the primers do not share sequence homology with human coronavirus 229E; human coronavirus OC43; influenza virus type A; influenza virus type B; parainfluenza virus types 1, 2 or 3; respiratory syncytial virus type A, respiratory syncytial virus type B; or human metapneumovirus.   
     
     
         2 . A method for detecting SARS-COV, which comprises amplifying a nucleic acid sample obtained from an individual by PCR using the primer set of  claim 1 . 
     
     
         3 . The method of  claim 2 , wherein the PCR is performed in a polypropylene tube, a 96-well plate, or a silicon-based micro PCR chip. 
     
     
         4 . The method of  claim 2 , wherein the PCR is a two-step PCR comprising a denaturation step and an annealing and extension step or a three-step PCR comprising a denaturation step, an annealing step, and an extension step. 
     
     
         5 . The method of  claim 2 , wherein the PCR is performed using 0.2-1 μM of each primer and 0.01 pg to 1 μg of a template DNA. 
     
     
         6 . The method of  claim 2 , wherein a time required for the PCR on a silicon-based micro PCR chip is 30 minutes or less. 
     
     
         7 . The method of  claim 2 , wherein the PCR is performed in three-step thermal cycling conditions of denaturation at 86-97° C. for 1-30 seconds, annealing at 50-70° C. for 1-30 seconds, and extension at 60-72° C. for 1-30 seconds, or in two-step thermal cycling conditions of denaturation at 86-97° C. for 1-30 seconds and annealing and extension at 50-70° C. for 5-30 seconds. 
     
     
         8 . The method of  claim 6 , wherein the silicon-based micro PCR chip comprises:
 a silicon substrate, a surface of which is formed with a PCR chamber made by silicon lithography and the other surface is formed with a heater for heating the PCR chamber; and   a glass substrate having an inlet and an outlet.   
     
     
         9 . A SARS-COV detection kit comprising the primer set of  claim 1 . 
     
     
         10 . The primer set of  claim 1 , wherein the primers do not share sequence homology with each of human coronavirus 229E, human coronavirus OC43, influenza virus type A, influenza virus type B; parainfluenza virus types 1, 2, and 3; respiratory syncytial virus types A and B; and human metapneumovirus.