Novel oligonucleotide arrays and their use for sorting, isolating, sequencing, and manipulating nucleic acids

Abstract

The invention provides hyperactive mutant recombinases and hybrid mutant recombinases, and methods for their identification. Also provided are nucleic acids encoding hyperactive mutant recombinases and hybrid recombinases, as well as vectors and host cells. Host cells include eukaryotic cells capable of expressing said recombinases and carrying out site-specific recombination in the cell. The mutant recombinases may be used, for example, in biotechnology, gene therapy or transgenic applications.

Description

FIELD OF THE INVENTION [0001] This invention is in the field of sorting, isolating, sequencing, and manipulating nucleic acids. BACKGROUND OF THE INVENTION [0002] Ordered arrays of oligonucleotides immobilized on a solid support have been proposed for sequencing DNA fragments. It has been recognized that hybridization of a cloned single-stranded DNA fragment to all possible oligonucleotide probes of a given length can identify the corresponding, complementary oligonucleotide segments that are present somewhere in the fragment, and that this information can sometimes be used to determine the DNA sequence. Use of arrays can greatly facilitate the surveying of a DNA fragment's oligonucleotide segments. There are two approaches currently being employed. [0003] In one approach, each oligonucleotide probe is immobilized on a solid support at a different predetermined position, forming an array of oligonucleotides. The array allows one to simultaneously survey all the oligonucleotide segme...

AI Technical Summary

Benefits of technology

"The invention is about new methods and arrays for analyzing and sorting nucleic acid strands. These methods involve using binary arrays with immobilized oligonucleotides that can be tailored to specific sequences. These arrays can be used to sort strands based on their terminal sequences or internal sequences. The methods also involve using sectioned arrays, which allow for separate reactions in each section and can be used to isolate individual strands from complex mixtures. The invention has several technical effects, including increased specificity and sensitivity in hybridization, the ability to sort strands based on their internal sequences, and the ability to isolate individual strands from complex mixtures."

Problems solved by technology

There is an inherent redundancy in the data, due to the overlapping nature of the oligonucleotides.

There is, however, an important limitation to sequencing by known surveying techniques.

When this occurs, the sequence of the DNA cannot be unambiguously determined.

The longer the DNA sequence, the worse this problem becomes.

However, the completion of a clone library is essentially an asymptotic process.

Moreover, there is no way to know whether the library is comprehensive or not, until the sequenced fragments are finally assembled.

The cloning of fragments of an entire genome is extremely slow and tedious.

This latter circumstance makes PCR, in its current form, barely useful for the preparation of individual fragments of unknown nucleotide sequences.

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