Method for generating a subtracted cDNA library and uses of the generated library
AssigneeThe Trustees of Columbia University in the City of New York
InventorsPaul B. Fisher, Hongping Jiang
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Abstract
This invention provides a method of generating a subtracted cDNA library of a cell comprising: a) generating a cDNA library of the cell; b) isolating double-stranded DNAs from the cDNA library; c) releasing the double-stranded cDNA inserts from the double-stranded DNAs; d) denaturing the isolated double-stranded cDNA inserts; e) hybridizing the denatured double-stranded cDNA inserts with a labelled single-stranded nucleic acid molecules which are to be subtracted from the cDNA library; and f) separating the hybridized labeled single-stranded nucleic acid molecule from the double-stranded cDNA inserts, thereby generating a subtracted cDNA library of a cell.
Claims22
A method of generating a subtracted cDNA library of a population of cells comprising:
- a) generating a .lambda. ZAP cDNA library of the cells;
- b) converting the .lambda. ZAP cDNA library into a phagemid cDNA library;
- c) preparing double-stranded phagemid DNAs;
- d) isolating double-stranded DNAs from the prepared phagemid DNAs;
- e) releasing the double-stranded cDNA inserts from the double-stranded DNAs by digestion of the nucleic acids with appropriate restriction enzyme;
- f) denaturing the isolated double-stranded cDNA inserts;
- g) hybridizing the denatured double-stranded cDNA inserts with labelled single stranded nucleic acid molecules, which are to be subtracted from the cDNA library;
- and
- h) separating the hybridized labelled single-stranded nucleic acid molecules from the double stranded cDNA inserts, thereby generating a subtracted cDNA library of a population of cells.
Dependencies are inferred from the claim text.