Method for detecting transcription templates

Abstract

Methods are provided for detecting the sense and antisense transcripts and for determining template strand of a genomic DNA. Exemplary methods include reverse transcribing transcripts without second strand synthesis. The resulting single stranded stranded DNA is labeled and detected using nucleic acid probe arrays. In a particularly preferred embodiment, actinomycin is used to inhibit the synthesis of second strand cDNA during reverse transcription.

1 . A method for detecting a plurality of transcripts comprising: 
 synthesizing a plurality of cDNAs complementary with the transcripts by reverse transcription; wherein the synthesis of second strand cDNA is inhibited; and    hybridizing the cDNAs or nucleic acids derived from the cDNAs with a nucleic acid probe array to detect the transcripts.    
     
     
         2 . The method of  claim 1  wherein the synthesis of the second strand cDNA is inhibited by the presence of actinomycin.  
     
     
         3 . The method of  claim 2  wherein the cDNAs or nucleic acids derived from the cDNAs are labeled.  
     
     
         4 . The method of  claim 2  wherein the nucleic acid probe array is an oligonucleotide probe array.  
     
     
         5 . The method of  claim 4  wherein the nucleic acid probe array has at least 400 probes per cm 2 .  
     
     
         6 . The method of  claim 5  wherein the nucleic acid probe array has at least 1000 probes per cm 2 .  
     
     
         7 . The method of  claim 6  wherein the nucleic acid probe array has at least 10000 probes per cm 2 .  
     
     
         8 . The method of  claim 4  wherein the nucleic acid probe array comprises at least least one probe against a target sequence and one probe against the reverse complementary sequence of the target sequence.  
     
     
         9 . The method of  claim 8  wherein the nucleic acid probe array comprises at least least 100 probes against at least 100 target sequences and at least 100 probes against at least 100 reverse complementary sequences of the target sequences.  
     
     
         10 . The method of  claim 9  wherein the nucleic acid probe array comprises at least 1000 probes against at least 1000 target sequences and at least 1000 probes against at least 1000 reverse complementary sequences of the target sequences.  
     
     
         11 . The method of  claim 10  wherein the nucleic acid probe array comprises at least 3000 probes against at least 3000 target sequences and at least 3000 probes against at least 3000 reverse complementary sequences of the target sequences.  
     
     
         12 . A method for detecting transcribed regions of a genome comprising 
 obtaining a sample comprising transcripts transcribed from the genome;    synthesizing single stranded cDNAs complementary with the transcripts, wherein the synthesis of second strand cDNA is inhibited; and    hybridizing the cDNAs or nucleic acids derived from the cDNAs with a nucleic acid probe array, wherein the nucleic acid probe array has probes targeting both strands of the genomic DNA in interested regions.    
     
     
         13 . The method of  claim 12  wherein the synthesis of the second strand cDNA is inhibited by the presence of actinomycin.  
     
     
         14 . The method of  claim 13  wherein the cDNAs or nucleic acids derived from the cDNAs are labeled.  
     
     
         15 . The method of  claim 14  wherein the nucleic acid probe array is an oligonucleotide probe array.  
     
     
         16 . The method of  claim 15  wherein the nucleic acid probe array has at least 400 probes per cm 2 .  
     
     
         17 . The method of  claim 16  wherein the nucleic acid probe array has at least 1000 probes per cm 2 .  
     
     
         18 . The method of  claim 17  wherein the nucleic acid probe array has at least 10000 probes per cm 2 .  
     
     
         19 . The method of  claim 1   2  further comprising determining the template strand for at least one transcript, and wherein the probe array contains probes against both strand of the genomic DNA region where the transcript is transcribed.  
     
     
         20 . An assay kit comprising: 
 reagents necessary for a reverse transcription reaction;    an inhibitor of second strand cDNA synthesis and    a nucleic acid probe array.    
     
     
         21 . The kit of  claim 20  wherein the inhibitor is actinomycin D.  
     
     
         22 . The kit of  claim 21  wherein the nucleic acid probe array is an oligonucleotide probe array.  
     
     
         23 . The kit of  claim 22  wherein the nucleic acid probe array has at least 400 probes per cm 2 .  
     
     
         24 . The kit of  claim 23  wherein the nucleic acid probe array has at least 1000 probes per cm 2 .  
     
     
         25 . The kit of  claim 24  wherein the nucleic acid probe array has at least 10000 probes per cm 2 .  
     
     
         26 . The kit of  claim 25  wherein the nucleic acid probe array comprises at least one probe against a target sequence and one probe against the reverse complementary sequence of the target sequence.  
     
     
         27 . The kit of  claim 26  wherein the nucleic acid probe array comprises at least 100 probes against at least 100 target sequences and at least 100 probes against at least 100 reverse complementary sequences of the target sequences.  
     
     
         28 . The kit of  claim 27  wherein the nucleic acid probe array comprises at least 1000 probes against at least 1000 target sequences and at least 1000 probes against at least 1000 reverse complementary sequences of the target sequences.  
     
     
         29 . The kit of  claim 28  wherein the nucleic acid probe array comprises at least 3000 probes against at least 3000 target sequences and at least 3000 probes against at least 3000 reverse complementary sequences of the target sequences.