The present invention pertains to a method to isolate, separate, enrich or amplify a targeted
nucleotide polymer such as mRNA through selective reverse transcription of the targeted
polymer into cDNA from a sample comprising of chemically identical or similar
polynucleotide polymers such as rRNA. The enrichment of the targeted
nucleic acid such as mRNA is accomplished by blocking the reverse transcription of undesired rRNA while allowing unrestricted reverse transcription of the targeted
polymer. The invention also embodies that the cleavage of the non-targeted
nucleic acid such as rRNA bound to an
oligonucleotide through enzymatic activity (
RNase H). The invention further embodies methods and kits to accomplish the utility of the invention through the following steps 1) 3′ tailing of chemically identical or similar
nucleotide polymers in a sample that includes bacterial mRNA 2) a 3′
tail capable of binding to a oligo-dN primer 3) at least one
oligonucleotide capable of preventing the extension of oligo-dN bound to at least one non-targeted
nucleotide polymers by
a DNA polymerase such as a
reverse transcriptase without restricting conversion of bacterial mRNA into cDNA 4) where the non-targeted molecule is prevented as a template for cDNA synthesis by enzymatic cleavage (
RNase H) of template (rRNA)-
oligonucleotide hybrid 5) where the
reverse transcriptase is physically blocked by the oligonucleotide bound to the non-targeted nucleic acids such as rRNA 5) purification of the selectively transcribed cDNA. In further embodiments of the present invention, methods and composition to enable the study of bacterial transcriptomics-an analysis of genes expressed by a bacterial infection of a host, an isolated bacterial culture or a bacterial
community, such as recovered from soil, intestine, mouth,
biofilm, water etc are also included for use in
DNA-
chip or sequencing analyses.