Construction of uni-directionally cloned cDNA libraries from messenger RNA for improved 3′ end DNA sequencing

A method of producing uni-directionally cloned complementary DNA libraries from mRNA, the method comprising:

• contacting mRNA with a primer mixture comprising a plurality of primers having different sequences, wherein each primer in the mixture comprises:
• at least 10 contiguous deoxythymidines in the 3′ end of the primer;
• at least two non-deoxythymidine nucleotides 3′ of the contiguous deoxythymidines;
• and
• a restriction enzyme sequence, which restriction enzyme sequence can be cleaved by a restriction enzyme when the primer is paired with its complement to provide a double-stranded restriction enzyme sequence;
• reverse transcribing the mRNA to produce a DNA strand complementary to the mRNA;
• substantially removing the polyT tail from the DNA strand;
• inserting the DNA strand uni-directionally into a cloning vector, and amplifying the DNA strand using a DNA polymerase;
• wherein a uni-directionally cloned complementary DNA library is produced.