Bioo Scientific Corporation

Kevin Allen

Adam R Morris

Masoud Toloue

Suk Ho Eun

Kerry Gunning

Oligonucleotide primers and methods of use in producing sequencing libraries are provided according to aspects of the present disclosure which include, from 5' to 3′, a homopolymer-hybridizing region, and an anchor region comprising 5′-(λ)N-3′, wherein the homopolymer-hybridizing region is a contiguous sequence of 5 to 20 elements, wherein the elements are nucleotides or Tincreasing nucleotide analogs, wherein at least 4 of the elements are Tincreasing nucleotide analogs, wherein the homopolymer-hybridizing region hybridizes to a complementary homopolymer tract of a target nucleic acid, wherein the complementary homopolymer tract comprises a contiguous sequence of complementary elements, wherein X is any nucleotide or nucleotide analog with the proviso that λ is not a nucleotide or nucleotide analog complementary to a complementary element of the complementary homopolymer tract, and wherein N is any nucleotide or nucleotide analog.

Compositions and methods for generating massively parallel nucleic acid sequencing libraries

This disclosure provides, among other things, a method for making a cDNA library. In some embodiments the method may comprise reverse transcribing mRNA to produce DNA:mRNA hybrids, treating the DNA:mRNA hybrids with RNAseH to produce mRNA fragments, and reverse transcribing the mRNA fragments.

Method for making a cDNA library

This disclosure provides, among other things, a 5' adapter of the formula 3&#x2032;*&#x2014;X&#x2014;(5&#x2032;5&#x2032;)&#x2014;Y&#x2014;3&#x2032;, where: 3&#x2032;* is a blocked 3&#x2032; end, X is a synthetic sequence, (5&#x2032;5&#x2032;) is an internal 5&#x2032;-5&#x2032; linkage, Y is an adapter sequence, and 3&#x2032; is a hydroxylated 3&#x2032; end. In use, sequence X hybridizes to sequence X&#x2032; in a population of RNA molecules of formula R&#x2014;X&#x2032;, which increases the efficiency of ligation of the 5&#x2032; adapter to the nucleic acid molecules.

5' adapter comprising an internal 5&#x2032;-5&#x2032; linkage

This disclosure provides, among other things, a method for making a cDNA library. In some embodiments the method may comprise adding a polyA tail to the longer RNA fragments but not the shorter RNA fragments in a sample by incubating the population of RNA fragments with a polyA polymerase, wherein the reaction conditions used preferentially tail only the longer fragments but not the shorter fragments.

Size selection of RNA using poly(A) polymerase

The current teachings relate to methods for reducing adapter-dimer formation, particularly when preparing nucleic acids of interest for subsequent amplification and/or sequencing. Also described are kits for use in performing certain disclosed methods.

Growing community of inventors

Austin, TX, United States of America

Kevin Allen