Allelic frequencies and statistical data obtained from 12 codis STR loci in an admixed population of the Brazilian Amazon

The allelic frequencies of 12 short tandem repeat loci were obtained from a sample of 307 unrelated individuals living in Macapá, a city in the northern Amazon region, Brazil. These loci are the most commonly used in forensics and paternity testing. Based on the allele frequency obtained for the population of Macapá, we estimated an interethnic admixture for the three parental groups (European, Native American and African) of, respectively, 46%, 35% and 19%. Comparing these allele frequencies with those of other Brazilian populations and of the Iberian Peninsula population, no significant distances were observed. The interpopulation genetic distances (F(ST) coefficients) to the present database ranged from F(ST) = 0.0016 between Macapá and Belém to F(ST) = 0.0036 between Macapá and the Iberian Peninsula.     

Inter- and intra-specific variation among five Erythroxylum taxa assessed by AFLP

BACKGROUND: and Aims The four cultivated Erythroxylum taxa (E. coca var. coca, E. novogranatense var. novogranatense, E. coca var. ipadu and E. novogranatense var. truxillense) are indigenous to the Andean region of South America and have been cultivated for folk-medicine and, within the last century, for illicit cocaine production. The objective of this research was to assess the structure of genetic diversity within and among the four cultivated alkaloid-bearing taxa of Erythroxylum in the living collection at Beltsville Agricultural Research Center. METHODS: Amplified fragment length polymorphism (AFLP) fingerprinting was performed in 86 Erythroxylum accessions using a capillary genotyping system. Cluster analysis, multidimensional scaling (MDS) and analysis of molecular variance (AMOVA) were used to assess the pattern and level of genetic variation among and within the taxa. KEY RESULTS: A clear distinction was revealed between E. coca and E. novogranatense. At the intra-specific level, significant differentiation was observed between E. c. var. coca and E. c. var. ipadu, but the differentiation between E. n. var. novogranatense and E. n. var. truxillense was negligible. Erythroxylum c. var. ipadu had a significantly lower amount of diversity than the E. c. var. coca and is genetically different from the E. c. var. ipadu currently under cultivation in Colombia, South America. CONCLUSIONS: There is a heterogeneous genetic structure among the cultivated Erythroxylum taxa where E. coca and E. novogranatense are two independent species. Erythroxylum coca var. coca is most likely the ancestral taxon of E. c. var. ipadu and a founder effect may have occurred as E. c. var. ipadu moved from the eastern Andes in Peru and Bolivia into the lowland Amazonian basin. There is an indication of artificial hybridization in coca grown in Colombia.     

A cell-based cGMP assay useful for ultra-high-throughput screening and identification of modulators of the nitric oxide/cGMP pathway

We have established a rapid, homogeneous, cell-based, and highly sensitive assay for guanosine 3'-5'-cyclic monophosphate (cGMP) that is suitable for fully automated ultra-high-throughput screening. In this assay system, cGMP production is monitored in living cells via Ca2+ influx through the olfactory cyclic nucleotide-gated cation channel CNGA2, acting as the intracellular cGMP sensor. A stably transfected Chinese hamster ovary (CHO) cell line was generated recombinantly expressing soluble guanylate cyclase, CNGA2, and aequorin as a luminescence indicator for the intracellular calcium concentration. This cell line was used to screen more than 900,000 compounds in an automated ultra-high-throughput screening assay using 1536-well microtiter plates. In this way, we have been able to identify BAY 58-2667, a member of a new class of amino dicarboxylic acids that directly activate soluble guanylate cyclase. The assay system allows the real-time cGMP detection within living cells and makes it possible to screen for activators and inhibitors of enzymes involved in the nitric oxide/cGMP pathway.     

Fractionation of serum components using nanoporous substrates

Numerous previously uncharacterized molecules resident within the low molecular weight circulatory proteome may provide a picture of the ongoing pathophysiology of an organism. Recently, proteomic signatures composed of low molecular weight molecules have been identified using mass spectrometry combined with bioinformatic algorithms. Attempts to sequence and identify the molecules that underpin the fingerprints are currently underway. The finding that many of these low molecular weight molecules may exist bound to circulating carrier proteins affords a new opportunity for fractionation and separation techniques prior to mass spectrometry-based analysis. In this study we demonstrate a method whereby nanoporous substrates may be used for the facile and reproducible fractionation and selective binding of the serum-based biomarker material, including subcellular proteins found within the serum. Aminopropyl-coated nanoporous silicon, when exposed to serum, can deplete serum of proteins and yield a serum with a distinct, altered MS profile. Additionally, aminopropyl-coated, nanoporous controlled-pore glass beads are able to bind a subset of serum proteins and release them with stringent elution. The eluted proteins have distinct MS profiles, gel electrophoresis profiles, and differential peptide sequence identities, which vary based on the size of the nanopores. These material surfaces could be employed in strategies for the harvesting and preservation of labile and carrier-protein-bound molecules in the blood.     

Palindrome-mediated chromosomal translocations in humans

Recently, it has emerged that palindrome-mediated genomic instability contributes to a diverse group of genomic rearrangements including translocations, deletions, and amplifications. One of the best studied examples is the recurrent t(11;22) constitutional translocation in humans that has been well documented to be mediated by palindromic AT-rich repeats (PATRRs) on chromosomes 11q23 and 22q11. De novo examples of the translocation are detected at a high frequency in sperm samples from normal healthy males, but not in lymphoblasts or fibroblasts. Cloned breakpoint sequences preferentially form a cruciform configuration in vitro. Analysis of the junction fragments implicates frequent double-strand-breaks (DSBs) at the center of both palindromic regions, followed by repair through the non-homologous end joining (NHEJ) pathway. We propose that the PATRR adopts a cruciform structure in male meiotic cells, creating genomic instability that leads to the recurrent translocation.     

Nanoporous surfaces as harvesting agents for mass spectrometric analysis of peptides in human plasma

Silica-based nanoporous surfaces have been developed in order to capture low molecular weight peptides from human plasma. Harvested peptides were subjected to mass spectrometric analysis by using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) as a means of detecting and assessing the bound molecules. Peptide profiles consisting of about 70 peaks in the range 800-10,000 m/z were generated. The method could allow detection of small peptides at ng/mL concentration levels, either in standard solutions or in plasma. The same molecular cutoff effect was observed for mixtures of standard proteins and peptides incubated with silicon-based nanoporous surfaces.