Qiagen Gmbh

Wolfgang Krebs

Nan Fang

Isabell Czolkos

The invention relates to a method for preparing a strand-specific library from an nucleic acid or preferably RNA sample, for RNA comprising the steps of: (i) optionally fragmenting said RNA sample, (ii) generating a plurality of first cDNA strands by subjecting said fragmented RNA to reverse transcription by using a reverse transcriptase and first oligonucleotide primers, (iii) generating a plurality of second cDNA strands by using a DNA polymerase, second oligonucleotide primers, and the plurality of first cDNA strands, and (iv) ligating adapters to the 3' and 5&#x2032; termini of the of double-stranded cDNA, (v) wherein the first cDNA strand allows no adapter ligation at its 5&#x2032; terminus and said second cDNA strand allows adapter ligation at its 5&#x2032; terminus, or vice versa, and, (v) optionally cloning, sequencing or otherwise using the strand-specific library. The invention also relates to a kit for preparing a strand-specific library from an RNA sample, wherein said kit comprises, (i) random oligonucleotide primers comprising a 5&#x2032; terminus nucleotide which allows no adaptor ligation, (ii) random oligonucleotide primers comprising a 5&#x2032; terminus nucleotide which allows adaptor ligation, (iii) optionally a reverse transcriptase for synthesizing a first cDNA strand complementary to the template RNA, (iv) optionally dNTPs, and (v) optionally a DNA polymerase.

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Hilden, Germany

Wolfgang Krebs