Abstract: Compositions and methods are provided for creating and identifying mutant carbohydrate-binding proteins that reversibly bind to carbohydrate substrates under conditions where the native protein remains bound. Examples of modified chitin-binding domains are provided which can be eluted from chitin in the presence of a reducing agent or at a pH within the range of 5-10.

Abstract: Specified restriction endonucleases have been characterized for the first time by their amino acid and DNA sequences. These sequences and those with at least 90% identity thereto have been used as probes in sequence similarity analyses to identify sequence matches in a sequence database that corresponds to novel restriction endonucleases or isoschizomers. The sequence similarity analyses includes selecting a positive sequence match from any sequence producing an expectation value of less than or equal to e-02.

Abstract: The invention relates to new proteins called alkylcytosine transferases (ACTs) derived from O6-alkylguanine-DNA alkyltransferase, and to substrates for ACTs specifically transferring a label to these ACTs and to fusion proteins comprising these. The substrates according of the invention are substituted cytosines of formula (I) wherein R1 is an aromatic or a heteroaromatic group, or an optionally substituted unsaturated alkyl, cycloalkyl or heterocyclyl group with the double bond connected to OCH2—; R2 is a linker; and L is a label or a plurality of same or different labels. The invention further relates to methods of transferring label L from these substrates of formula (I) to ACTs and ACT fusion proteins. The system of ACT-compound of formula (I) is particularly suitable for double labelling studies together with the known system O6-alkylguanine-DNA alkyltransferase (AGT)-benzylguanines.

Abstract: Methods and compositions are provided that relate to obtaining a recombinant DNA and RNA cleaving nuclease. This involves the over-expression of a fusion protein between maltose-binding protein and a truncated nuclease in a soluble form in the cytoplasm of a host cell from which it can be readily extracted.

Abstract: The invention relates to pyrimidines suitable as substrates for O6-alkylguanine-DNA alkyltransferases (AGT) of formula (I) wherein R1 is hydrogen, lower alkyl, halogen, cyano, trifluoromethyl or azido; R2 is a linker; and L is a label or a plurality of same or different labels. The invention further relates to methods of transferring a label from pyrimidines of formula (I) to O6-alkylguanine-DNA alkyltransferases (AGT) and AGT fusion proteins.

Abstract: The invention relates to substrates for O6-alkylguanine-DNA alkyltransferases (AGT) of formula R1-A-X—CH2—R3—R4-L1, wherein A is a group recognized by AGT as a substrate, X is oxygen or sulfur, R1 is a group —R2-L2 or a group R5, R2 and R4 are, independently of each other, a linker, R3 is an aromatic or a heteroaromatic group, or an optionally substituted unsaturated alkyl, cycloalkyl or heterocyclyl group with the double bond connected to CH2, R5 is arylmethyl or heteroarylmethyl or an optionally substituted cycloalkyl, cycloalkenyl or heterocyclyl group, L1 is a label, a plurality of same or different labels, a bond connecting R4 to A forming a cyclic substrate, or a further group —R3—CH2—X-A-R1, and L2 is a label or a plurality of same or different labels. The invention further relates to methods of transferring a label from these substrates to O6-alkylguanine-DNA alkyltransferases (AGT) and AGT fusion proteins.

Abstract: Methods and compositions are provided for repairing a polynucleotide so that it can be copied with improved fidelity and/or yield in, for example, an amplification reaction. This involves the use of a reaction mixture that includes a ligase and a cofactor selected from NAD+ or ATP and incubating the polynucleotide with the reaction mixture in the absence of Endonuclease VI. The reaction mixture may further contain an AP endonuclease and a polymerase. If used, these enzymes may be selected according to their ability to withstand high temperatures. For example, the reaction mixture may be used prior to a polynucleotide synthesis reaction in which case enzymes that are not thermophilic may be used. The repair reaction is not time sensitive with respect to seconds, minutes or hours of incubation in the enzyme mixture in as much as the repair is effected rapidly and prolonged incubation is not generally adverse.

Abstract: Compositions and methods are provided for selection and enrichment of a target gene from a library of polynucleotide sequences such as might be formed from a genome or by random mutagenesis of a genetic sequence. The selection and enrichment occurs in aqueous droplets formed in an emulsion that compartmentalize individual polynucleotides from the library or a plurality of polynucleotides that may include polynucleotides not derived from the library, transcription and translation reagents and optionally additional chemical and enzyme reagents. The selection and enrichment method utilizes a polynucleotide adaptor which when ligated to the polynucleotide fragment enables amplification to occur in the presence of an adaptor specific primer.

Abstract: Methods and compositions are described that relate to obtaining concentrated preparations of secreted recombinant proteins. These proteins are expressed in the form of fusion proteins with a chitin-binding domain (CBD). The fusion proteins are capable of being concentrated in the presence of chitin. Also described is: a shuttle vector that includes a modified LAC4 promoter; a chitinase-negative host cell; a CBD capable of eluting from chitin under non-denaturing conditions; and sterilized chitin, which can be optionally magnetized for facilitating recovery of recombinant protein.

Abstract: Methods are provided for making restriction endonucleases with reduced star activity by one or more targeted mutations to a catalytic site within the restriction endonuclease. Examples of modifications to restriction endonucleases with significant sequence identity with KpnI are provided and reduced star activity demonstrated.

Abstract: The present invention relates to compositions including: (1) isolated DNA encoding the StuI restriction endonuclease and isolated DNA encoding cognate and non-cognate methylase; (2) vectors and cells containing the isolated DNA; and (3) methods for producing the StuI restriction endonuclease.

Abstract: Compositions and methods are provided that relate to a modified DNA cleaving enzyme having at least 35% amino acid sequence identity with T7 Endo I. The modified enzyme includes two catalytic centers separated by a ?-bridge where the ?-bridge contains at least one mutation having an effect of altering enzyme cleavage activity compared to the unmodified enzyme. Activities associated with the modified DNA cleaving enzyme that can be modulated in different reaction conditions include at least one of: (a) non-sequence specific nicking activity; (b) cleaving the second strand of a duplex DNA at a preexisting nick site to produce a linear duplex with a single strand overhang; (c) non-sequence specific DNA cleavage; (d) cleaving DNA flanking a mismatch; and (e) cleavage at a cruciform structure in a DNA duplex.