Abstract: Ordered assembly of large numbers of fragments into a single large DNA have been improved in both frequency and fidelity of the assembled product. This has been achieved by novel compositions and methods that are utilized in a computer system that integrates comprehensive ligation data from multiple sources to provide optimized synthetic overhangs or overhangs from restriction endonuclease cleavage on DNA fragments for assembly by ligation. Intragenic cut sites are avoided by the use of a novel restriction endonuclease which recognizes 7 nucleotides (bases) and cuts DNA to create 4-base overhangs with the help of a synthetic activator oligonucleotide. Variations in ligation preferences by different ligases provide extra precision in assembly reactions. The use of the improved methods are exemplified by the successful assembly from 52 fragments of a viral genome and also a 52 fragment ordered assembly of a bacteria operon.

Abstract: A composition and its uses and additionally a kit are provided. The composition is a synthetic self-complementary oligonucleotide that has a double-stranded region and a loop, wherein the double-stranded region contains a binding sequence for PaqCl. Additionally, the oligonucleotide includes unligatable 3? and 5? ends that cannot be cleaved by PaqCl. This oligonucleotide composition has been combined with PaqCl or a variant of PaqCl Type IIS restriction endonuclease in a reaction mixture, where the reaction mixture includes PaqCl or variant that can further be combined with a ligase and optionally a deadenylase, crowding molecule such as PEG and/or a repair enzyme such as Endo MS. The kit includes the oligonucleotide and Type IIS restriction endonuclease in the same or different containers.

Abstract: Methods and compositions are provided for optimizing ordered assembly of a plurality of polynucleotide fragments. The optimization involves providing sets of overhang sequences with preferred experimental conditions for high fidelity ordered assembly of polynucleotide fragments by ligation under selected experimental conditions. The methods and compositions provide the use of a computer system with inputs having a plurality of menus and outputs that include a variety of media interfaces. The computer system has access to a ligation frequency database to provide sets of overhang sequences for efficient joining of multiple fragments into the target nucleic acid.

Abstract: Mutant bacteriophage DNA ligases that have increased tolerance to salt and/or heat is provided. Methods, compositions and kits that employ the same are also provided.

Abstract: Compositions and methods are provided for ligating polynucleotides having a length that is greater than 8 nucleotides on an RNA splint. The ligation reaction provides consistent results in high or low ATP concentrations. The reaction can occur rapidly and is generally at least 10 fold more efficient than T4DNA ligase under optimal conditions for T4DNA ligase and the reaction time is less than 6 hours for example, less than 1 hour.

Abstract: Compositions and methods are provided for ligating polynucleotides having a length that is greater than 8 nucleotides on an RNA splint. The ligation reaction provides consistent results in high or low ATP concentrations. The reaction can occur rapidly and is generally at least 10 fold more efficient than T4DNA ligase under optimal conditions for T4DNA ligase and the reaction time is less than 6 hours for example, less than 1 hour.

Abstract: The present invention provides synthetic Moraxella catarrhalis lipooligosaccharide (LOS)-based oligosaccharides and conjugates containing various M. catarrhalis serotype-specific oligosaccharide antigens or various core M. catarrhalis oligosaccharide structures or motifs corresponding to one or more of the three major serotypes and/or members within a given serotype. The oligosaccharides may be synthesized by a chemical assembly methodology relying on a limited number of monosaccharide and disaccharide building blocks. The invention further provides M. catarrhalis LOS-based immunogenic and immunoprotective compositions and antibodies derived therefrom for diagnosing, treating, and preventing infections caused by M. catarrhalis.

Abstract: The present invention provides synthetic Moraxella catarrhalis lipooligosaccharide (LOS)-based oligosaccharides and conjugates containing various M. catarrhalis serotype-specific oligosaccharide antigens or various core M. catarrhalis oligosaccharide structures or motifs corresponding to one or more of the three major serotypes and/or members within a given serotype. The oligosaccharides may be synthesized by a chemical assembly methodology relying on a limited number of monosaccharide and disaccharide building blocks. The invention further provides M. catarrhalis LOS-based immunogenic and immunoprotective compositions and antibodies derived therefrom for diagnosing, treating, and preventing infections caused by M. catarrhalis.

Abstract: Compositions and methods are provided for ligating polynucleotides having a length that is greater than 8 nucleotides on an RNA splint. The ligation reaction provides consistent results in high or low ATP concentrations. The reaction can occur rapidly and is generally at least 10 fold more efficient than T4DNA ligase under optimal conditions for T4DNA ligase and the reaction time is less than 6 hours for example, less than 1 hour.

Abstract: Methods and compositions are provided for increasing the ligation yields of polynucleotides in vitro.

Abstract: Compositions and methods are provided for ligating polynucleotides having a length that is greater than 8 nucleotides on an RNA splint. The ligation reaction provides consistent results in high or low ATP concentrations. The reaction can occur rapidly and is generally at least 10 fold more efficient than T4DNA ligase under optimal conditions for T4DNA ligase and the reaction time is less than 6 hours for example, less than 1 hour.

Abstract: The present invention provides synthetic Moraxella catarrhalis lipooligosaccharide (LOS)-based oligosaccharides and conjugates containing various M. catarrhalis serotype-specific oligosaccharide antigens or various core M. catarrhalis oligosaccharide structures or motifs corresponding to one or more of the three major serotypes and/or members within a given serotype. The oligosaccharides may be synthesized by a chemical assembly methodology relying on a limited number of monosaccharide and disaccharide building blocks. The invention further provides M. catarrhalis LOS-based immunogenic and immunoprotective compositions and antibodies derived therefrom for diagnosing, treating, and preventing infections caused by M. catarrhalis.

Abstract: The present invention provides synthetic Moraxella catarrhalis lipooligosaccharide (LOS)-based oligosaccharides and conjugates containing various M. catarrhalis serotype-specific oligosaccharide antigens or various core M. catarrhalis oligosaccharide structures or motifs corresponding to one or more of the three major serotypes and/or members within a given serotype. The oligosaccharides may be synthesized by a chemical assembly methodology relying on a limited number of monosaccharide and disaccharide building blocks. The invention further provides M. catarrhalis LOS-based immunogenic and immuno-protective compositions and antibodies derived therefrom for diagnosing, treating, and preventing infections caused by M. catarrhalis.

Abstract: The present invention provides synthetic Moraxella catarrhalis lipooligosaccharide (LOS)-based oligosaccharides and conjugates containing various M. catarrhalis serotype-specific oligosaccharide antigens or various core M. catarrhalis oligosaccharide structures or motifs corresponding to one or more of the three major serotypes and/or members within a given serotype. The oligosaccharides may be synthesized by a chemical assembly methodology relying on a limited number of monosaccharide and disaccharide building blocks. The invention further provides M. catarrhalis LOS-based immunogenic and immuno-protective compositions and antibodies derived therefrom for diagnosing, treating, and preventing infections caused by M. catarrhalis.