As population structure can result in spurious associations, it has constrained the use of association studies in human and plant genetics. Association mapping, however, holds great promise if true signals of functional association can be separated from the vast number of false signals generated by population structure. We have developed a unified mixed-model approach to account for multiple levels of relatedness simultaneously as detected by random genetic markers. We applied this new approach to two samples: a family-based sample of 14 human families, for quantitative gene expression dissection, and a sample of 277 diverse maize inbred lines with complex familial relationships and population structure, for quantitative trait dissection. Our method demonstrates improved control of both type I and type II error rates over other methods. As this new method crosses the boundary between family-based and structured association samples, it provides a powerful complement to currently available methods for association mapping.

A unified mixed-model method for association mapping that accounts for multiple levels of relatedness

Mutations in LRRK2 Cause Autosomal-Dominant Parkinsonism with Pleomorphic Pathology

http://theory.bio.uu.nl/immbio/Marks91.pdf

By-passing immunization: Human antibodies from V-gene libraries displayed on phage

9-Fluorenylmethoxycarbonyl (Fmoc) amino acids were first used for solid phase peptide synthesis a little more than a decade ago. Since that time, Fmoc solid phase peptide synthesis methodology has been greatly enhanced by the introduction of a variety of solid supports, linkages, and side chain protecting groups, as well as by increased understanding of solvation conditions. These advances have led to many impressive syntheses, such as those of biologically active and isotopically labeled peptides and small proteins. The great variety of conditions under which Fmoc solid phase peptide synthesis may be carried out represents a truly "orthogonal" scheme, and thus offers many unique opportunities for bioorganic chemistry.

Solid phase peptide synthesis utilizing 9‐fluorenylmethoxycarbonyl amino acids

A Sequencing Method Based on Real-Time Pyrophosphate

Various 5'O-N-protected deoxynucleoside-3'-O-beta-cyanoethyl-N,N-dialkylamino-/N- morpholinophosphoramidites were prepared from beta-cyanoethyl monochlorophosphoramidites of N,N-dimethylamine, N,N-diisopropylamine and N-morpholine. These active deoxynucleoside phosphates have successfully been used for oligodeoxynucleotide synthesis on controlled pore glass as polymer support and are very suitable for automated DNA-synthesis due to their stability in solution. The intermediate dichloro-beta- cyanoethoxyphosphine can easily be prepared free from any PC1(3) contamination. The active monomers obtained from beta-cyanoethyl monochloro N,N- diisopropylaminophosphoramidites are favoured. Cleavage of the oligonucleotide chain from the polymer support, N-deacylation and deprotection of beta-cyanoethyl group from the phosphate triester moiety can be performed in one step with concentrated aqueous ammonia. Mixed oligodeoxynucleotides are characterized by the sequencing method of Maxam and Gilbert.

Polymer support oligonucleotide synthesis XVIII1.2): use of β-cyanoethyi-N,N-dialkylamino-/N-morpholino phosphoramidite of deoxynucleosides for the synthesis of DNA fragments simplifying deprotection and isolation of the final product

The invention describes a new method to sequence DNA. The improvements over the existing DNA sequencing technologies are high speed, high throughput, no electrophoresis and gel reading artifacts due to the complete absence of an electrophoretic step, and no costly reagents involving various substitutions with stable isotopes. The invention utilizes the Sanger sequencing strategy and assembles the sequence information by analysis of the nested fragments obtained by base-specific chain termination via their different molecular masses using mass spectrometry, as for example, MALDI or ES mass spectrometry. A further increase in throughput can be obtained by introducing mass-modifications in the oligonucleotide primer, chain-terminating nucleoside triphosphates and/or in the chain-elongating nucleoside triphosphates, as well as using integrated tag sequences which allow multiplexing by hybridization of tag specific probes with mass differentiated molecular weights.

Dna sequencing by mass spectrometry

The invention provides fast and highly accurate mass spectrometer based processes for detecting a particular nucleic acid sequence in a biological sample. Depending on the sequence to be detected, the processes can be used, for example, to diagnose a genetic disease or chromosomal abnormality; a predisposition to a disease or condition, infection by a pathogenic organism, or for determining identity or heredity.

DNA diagnostics based on mass spectrometry

The invention relates to a process for the preparation of oligonucleotides by the following steps: reaction of a nucleoside with a phosphine derivative, reaction of the nucleotide derivative thus obtained with a nucleoside bonded to a polymeric carrier, oxidation of the carrier-bound nucleoside-nucleotide thus obtained with formation of phosphotriester groups, blocking of free primary 5'-OH groups, elimination of a protective group from the terminal 5'-OH group, where appropriate single or multiple repetition of the abovementioned steps to introduce further nucleoside phosphate or oligonucleoside phosphate units, and cleavage of the nucleoside-carrier bond and, where appropriate, elimination of all protective groups present in the oligonucleoside phosphates. The phosphine derivative used is a compound of the general formula III ##STR1## in which X and L can react with OH groups of the sugar units in the oligonucleotides, and R 3 is a protective group which can be liberated by β-elimination.

Process for the preparation of oligonucleotides

Processes and kits for simultaneously amplifying and sequencing nucleic acid molecules, and performing high throughput DNA sequencing are described.

Hubert Köster

Papers

Polymer support oligonucleotide synthesis XVIII1.2): use of β-cyanoethyi-N,N-dialkylamino-/N-morpholino phosphoramidite of deoxynucleosides for the synthesis of DNA fragments simplifying deprotection and isolation of the final product

Dna sequencing by mass spectrometry

DNA diagnostics based on mass spectrometry

Process for the preparation of oligonucleotides

Process for direct sequencing during template amplification