Molecular characterisation of 10 Dutch properdin type I deficient families: mutation analysis and X-inactivation studies.

Properdin type I deficiency is characterised by complete absence of extracellular properdin, a positive regulator of the alternative pathway of complement activation. Properdin deficiency is associated with increased susceptibility to severe meningococcal disease. We have identified the genetic defect in 10 Dutch families. Six different mutations and one sequence polymorphism in the properdin gene were found. All amino acid substitutions were limited to conserved amino acids in exons 7 and 8 in contrast to the premature stops that were found in other exons. The missense mutations may alter the protein conformation in such a way that properdin will not be secreted and therefore catabolised intracellularly. The decreased properdin levels found in some healthy females carrying one mutated properdin gene were studied for X-inactivation. Most carriers with extreme low or high properdin levels showed preferential X-inactivation for the normal or mutated X chromosome, respectively. We observed some exceptions, suggesting additional regulation of properdin excretion apart from X-inactivation.

Celsr1, a neural-specific gene encoding an unusual seven-pass transmembrane receptor, maps to mouse chromosome 15 and human chromosome 22qter.

We have identified Celsr1, a gene that encodes a developmentally regulated vertebrate seven-pass transmembrane protein. The extracellular domain of Celsr1 contains two regions each with homology to distinct classes of well-characterized motifs found in the extra-cellular domains of many cell surface molecules. The most N-terminal region contains a block of contiguous cadherin repeats, and C-terminal to this is a region containing seven epidermal growth factor-like repeats interrupted by two laminin A G-type repeats. Celsr1 is unique in that it contains this combination of repeats coupled to a seven-pass transmembrane domain. As part of the characterization of the Celsr1 gene, we have determined its chromosomal map location in both mouse and human. The European Collaborative Interspecific Backcross (EUCIB) and BXD recombinant inbred strains were used for mapping Celsr1 cDNA clones in the mouse, and fluorescence in situ hybridization was used to map human Celsr1 cosmid clones on metaphase chromosomes. We report that Celsr1 maps to proximal mouse Chromosome 15 and human chromosome 22qter, a region of conserved synteny. Reverse transcriptase-polymerase chain reaction analysis and in situ hybridization were used to determine the spatial restriction of Celsr1 transcripts in adult and embryonic mice. The results presented here extend our previous finding of expression of the Celsr1 receptor in the embryo and show that expression continues into adult life when expression in the brain is localized principally in the ependymal cell layer, choroid plexus, and the area postrema.

An integrated physical map of 210 markers assigned to the short arm of human chromosome 11.

Using a panel of patient cell lines with chromosomal breakpoints, we constructed a physical map for the short arm of human chromosome 11. We focused on 11p15, a chromosome band harboring at least 25 known genes and associated with the Beckwith-Wiedemann syndrome, several childhood tumors, and genomic imprinting. This underlines the need for a physical map for this region. We divided the short arm of chromosome 11 into 18 breakpoint regions, and a large series of new and previously described genes and markers was mapped within these intervals using fluorescence in situ hybridization. Cosmid fingerprint analysis showed that 19 of these markers were included in cosmid contigs. A detailed 10-Mb pulsed-field physical map of the region 11p15.3-pter was constructed. These three different approaches enabled the high-resolution mapping of 210 markers, including 22 known genes.

Separation and identification of illicit heroin samples by liquid chromatography using an alumina and C18 coupled column system and photodiode array detection.

The analysis of illicit heroin and opium samples on a coupled alumina and C18 column system is described. The compounds to be analysed can be divided into two groups: those with low pKa values, such as caffeine, papaverine and noscapine, and those with high pKa values, such as heroin, acetylcodeine, O6-monoacetylmorphine, procaine, codeine, morphine and strychnine. The first group can best be separated on a C18 column, whereas alumina is more suitable for the second group. Previously reported criteria for choosing proper buffer systems for ion-exchange separations on alumina were used together with an iterative regressive optimization procedure developed in our laboratory. The system can be used with and without valve-switching, depending on the sample type. The peak purity of the judicially important components heroin and O6-monoacetylmorphine has been checked with a photodiode array detector and by use of advanced software.