Complex high-resolution linkage disequilibrium and haplotype patterns of single-nucleotide polymorphisms in 2.5 Mb of sequence on human chromosome 21.

One approach to identify potentially important segments of the human genome is to search for DNA regions with nonrandom patterns of human sequence variation. Previous studies have investigated these patterns primarily in and around candidate gene regions. Here, we determined patterns of DNA sequence variation in 2.5 Mb of finished sequence from five regions on human chromosome 21. By sequencing 13 individual chromosomes, we identified 1460 single-nucleotide polymorphisms (SNPs) and obtained unambiguous haplotypes for all chromosomes. For all five chromosomal regions, we observed segments with high linkage disequilibrium (LD), extending from 1.7 to>81 kb (average 21.7 kb), disrupted by segments of similar or larger size with no significant LD between SNPs. At least 25% of the contig sequences consisted of segments with high LD between SNPs. Each of these segments was characterized by a restricted number of observed haplotypes,with the major haplotype found in over 60% of all chromosomes. In contrast, the interspersed segments with low LD showed significantly more haplotype patterns. The position and extent of the segments of high LD with restricted haplotype variability did not coincide with the location of coding sequences. Our results indicate that LD and haplotype patterns need to be investigated with closely spaced SNPs throughout the human genome, independent of the location of coding sequences, to reliably identify regions with significant LD useful for disease association studies.

A high-resolution radiation hybrid map of the human genome draft sequence.

We have constructed a physical map of the human genome by using a panel of 90 whole-genome radiation hybrids (the TNG panel) in conjunction with 40,322 sequence-tagged sites (STSs) derived from random genomic sequences as well as expressed sequences. Of 36,678 STSs on the TNG radiation hybrid map, only 3604 (9.8%) were absent from the unassembled draft sequence of the human genome. Of 20,030 STSs ordered on the TNG map as well as the assembled human genome draft sequence and the Celera assembled human genome sequence, 36% of the STSs had a discrepant order between the working draft sequence and the Celera sequence. The TNG map order was identical to one of the two sequence orders in 60% of these discrepant cases.

Functional loss of the horizontal doll's eye reflex following unilateral vestibular lesions.

The doll's eye reflex represents the vestibulo-ocular reflex (VOR) elicited by high-acceleration head rotation. After complete unilateral vestibular lesions, the ipsilateral, horizontal doll's eye reflex is replaced by a series of "catch-up" saccades. These cause permanent symptoms of blurred vision and dizziness during ipsilateral turns. We compared normal controls and patients with complete surgical lesions or canal paresis of up to 9 years duration via electronystagmography (ENG) to determine the usefulness of the doll's eye test as a diagnostic test for complete vestibular lesions. This test was found to be more sensitive in diagnosis of such lesions than head-shaking nystagmus, rotatory directional preponderance, and spontaneous nystagmus. It is also useful to document VOR function in patients in whom caloric irrigation is contraindicated.

Binding analysis of the estrogen receptor to its specific DNA target site in human breast cancer.

The estrogen receptor (ER) is a nuclear protein with a hormone- and a DNA-binding domain. We examined the DNA binding of ER in MCF-7 cells and 79 primary breast cancers by gel mobility shift assay using as a probe the estrogen response element (ERE). The mobility shift assay showed saturable, specific binding of ER to ERE in crude, high molar extracts containing greater than or equal to 4 mg/ml protein. Nonspecific binding was reduced by increasing concentrations of poly(deoxyinosidylate.deoxycytidylate) and shortening of the ERE probe from 35 to 15 base pairs. In the presence of Mg2+ the ER-ERE complex formation was hormone dependent at 22 degrees C but not at 37 degrees C. In the absence of Mg2+ estradiol was not necessary for ER-ERE complex formation. Correlation of the mobility shift assay with the hormone-binding (E2) assay showed agreement in 55 of the 79 tumors. Both assays were positive (E2 +/ERE+) in 35 cases and both were negative (E2-/ERE-) in 20 cases. In 11 tumors the hormone-binding assay was positive and the mobility shift assay negative (E2+/ERE-), suggesting an alteration of the DNA-binding domain. In 13 cancers the hormone-binding assay was negative and the mobility shift assay positive (E2-/ERE+) suggesting an alteration of the hormone-binding domain. By performing both hormone- and DNA-binding assays of ER and the hormone-binding assay of progesterone receptor (PR), we found the following subgroups of breast cancer: (a) E2+/ERE+/PR+, (b) E2+/ERE+/PR-, (c) E2+/ERE-/PR+, (d) E2+/ERE-/PR-, (e) E2-/ERE+/PR+, (f) E2-/ERE+/PR-, (g) E2-/ERE-/PR-. The simultaneous determination of 17 beta-estradiol and ERE binding may provide a better definition of the ER status of individual tumors and prove useful in refining endocrine therapy of patients with breast cancer.

A spontaneously produced lipopolysaccharide biosynthetic defect which causes both pleiotropic phage resistance and mucoid colony morphology in Salmonella anatum.

A spontaneously derived mutant of the smooth bacterial strain, Salmonella anatum A1, specifically blocks the DNA ejection function of bacteriophage E15 during infection. The mutant, AE15R-5, exhibits mucoid colony morphology but no evidence of colanic acid biosynthesis. It is resistant not only to bacteriophage E15, but also to all other smooth- and rough-specific phages which have been tested. Chemical, immunological and gel electrophoretic analyses indicate that its lipopolysaccharide (LPS) molecules fall into two categories: they are either highly truncated (probably heptoseless) or extremely large (complete LPS molecules with O-polysaccharides containing 80 or more repeat units). The antibiotic resistance pattern of AE15R-5 is roughly intermediate between that of a known heptoseless mutant, S. anatum MG4, and that of the parent strain, S. anatum A-1.