Exome Sequencing of a Multigenerational Human Pedigree

Over the next few years, the efficient use of next-generation sequencing (NGS) in human genetics research will depend heavily upon the effective mechanisms for the selective enrichment of genomic regions of interest. Recently, comprehensive exome capture arrays have become available for targeting approximately 33 Mb or ∼180,000 coding exons across the human genome. Selective genomic enrichment of the human exome offers an attractive option for new experimental designs aiming to quickly identify potential disease-associated genetic variants, especially in family-based studies. We have evaluated a 2.1 M feature human exome capture array on eight individuals from a three-generation family pedigree. We were able to cover up to 98% of the targeted bases at a long-read sequence read depth of ≥3, 86% at a read depth of ≥10, and over 50% of all targets were covered with ≥20 reads. We identified up to 14,284 SNPs and small indels per individual exome, with up to 1,679 of these representing putative novel polymorphisms. Applying the conservative genotype calling approach HCDiff, the average rate of detection of a variant allele based on Illumina 1 M BeadChips genotypes was 95.2% at ≥10x sequence. Further, we propose an advantageous genotype calling strategy for low covered targets that empirically determines cut-off thresholds at a given coverage depth based on existing genotype data. Application of this method was able to detect >99% of SNPs covered ≥8x. Our results offer guidance for “real-world” applications in human genetics and provide further evidence that microarray-based exome capture is an efficient and reliable method to enrich for chromosomal regions of interest in next-generation sequencing experiments

Crop Updates 2006 - Geraldton

This session covers twenty six papers from different authors 2006 Seasonal Outlook, David Stephens and Michael Meuleners, Department of Agriculture 2006 Wheat Market Outlook, Tony Smith, Plum Grove Commodity Trading Solutions Will Budgets Change in 2006? Peter Tozer, Department of Agriculture Wheat varieties – what does the industry need and how do we get closer? David Bowran Department of Agriculture Performance of Wheat Varieties in National Variety Testing (NVT) WA, Peter Burges, Agritech Crop Research Survey of lupin root health (in major production areas), Geoff Thomas, Bill MacLeod, Ken Adcock, Katie Bell, Ciara Beard and Anne Smith, Department of Agriculture Managing root disease under intensive cropping, Bill MacLeod, and Vivien Vanstone, Department of Agriculture Investigation into the adequacy OF SEALED FARM SILOS IN Western Australia to control phosphine-resistant Rhyzopertha dominica, C.R. Newman, Department of Agriculture Phosure – Extending the Life of Phosphine, Gabrielle Coupland and Ern Kostos, Cooperative Bulk Handling IWM performs over 5 years in 33 focus paddocks, Peter Newman and Glenn Adam, Department of Agriculture Maintaining wheat and lupin yields using phase pastures and shielded sprayers to manage increasing herbicide resistance, Caroline Peek, Nadine Eva, Chris Carter and Megan Abrahams, Department of Agriculture Can sheep selectively graze weeds out of crops? A model for using sheep rather than chemicals, Tim Wiley, Department of Agriculture and Dean Revell, CSIRO Livestock Environmental Protection (Clearing of Native Vegetation) Regulations 2004, Anne Finlay, Department of Environment What lies beneath? – Understanding constraints to productivity below the soil surface, Stephen Davies and Chris Gazey, Department of Agriculture, Bob Gilkes, Dan Evans and Tania Liaghati, University of Western Australia Phoma blight (P. schneiderae), a risk for WA lupins? Geoff Thomas and Mark Sweetingham, Department of Agriculture The 2005 Wheat streak mosaic virus epidemic in New South Wales and the threat posed to the Western Australian wheat industry, Roger Jones and Nichole Burges, Department of Agriculture Zone Management for fun and Profit, Department of Agriculture, Tony Rosser and Owen Mann, Great Northern Rural Annual Ryegrass Toxicity (ARGT) – How to manage the Risk, Marnie Thomas, Department of Agriculture The future of lupin varieties, Wayne Parker, Department of Agriculture Analysis of a wheat-pasture rotation in the 330mm annual rainfall zone using the STEP model, Andrew Blake and Caroline Peek, Department of Agriculture Value Added Opportunities for Lupins in High Value Feed and Food Markets, Jason Craig and Mark Tucek, Cooperative Bulk Handling An overview of the potential for a Biofuels Industry in Western Australia, Anne Wilkins and Nathan Hancock, Department of Agriculture The GMO Picture – a Reality Check, Dr Sue Sutherland, Department of Agriculture Nitrogen applied in splits rather than all applied at seeding returns higher gross income from wheat cropped on a leaching sandy soil at Muresk (Central agricultural region), Darshan Sharma, Department of Agriculture and Lionel Martin, Muresk Institute, Curtin University of Technology Potassium response in cereal cropping within the medium rainfall central wheatbelt, Jeff Russell, Department of Agriculture, Angie Roe and James Eyres Farm Focus consultants Western Region Barley Variety Guide2006, Alaina Smith, Blakely Paynter and Andrea Hills, Department of Agricultur

Solitomab, an EpCAM/CD3 bispecific antibody construct (BiTE), is highly active against primary uterine serous papillary carcinoma cell lines in vitro

BACKGROUND: Uterine serous carcinoma (USC) is an aggressive form of endometrial cancer which carries an extremely poor prognosis. Solitomab is a novel bispecific single-chain antibody construct which targets epithelial-cell-adhesion-molecule (EpCAM) on tumor cells and also contains a CD3 binding region. We evaluated the expression levels of EpCAM and the in vitro activity of solitomab against primary USC cell lines in vitro and ex-vivo in the ascites of USC patients. OBJECTIVES: To determine the frequency of expression of EpCAM on uterine serous carcinoma cell lines as well as the ability of solitomab to modulate immune responses (T-cell proliferation, activation, cytokine production, and tumor killing) to tumor cells when it is combined with lymphocytes and EpCAM positive cell lines or EpCAM positive acitic fluid in vitro. STUDY DESIGN: EpCAM expression was evaluated by flow cytometry in a total of 14 primary USC cell lines. Sensitivity to solitomab-dependent-cellular-cytotoxicity (ADCC) was tested against a panel of primary USC cell lines expressing different levels of EpCAM in standard 4h (51)Cr release-assays. The proliferative activity, activation, cytokine secretion (i.e., Type I vs Type II) and cytotoxicity of solitomab in autologous tumor-associated-T cells (TAL) in the ascitic fluid of USC patients was also evaluated by CFSE and flow-cytometry assays. Differences in EpCAM expression, ADCC levels were analyzed using upaired t test. T-cell activation marker increase and cytokine release were analyzed by paired t test. RESULTS: Surface expression of EpCAM was found in 85.7% (12 out of 14) of the USC cell lines tested by flow cytometry. EpCAM positive cell lines were found resistant to NK or T-cell-mediated killing after exposure to peripheral blood lymphocytes (PBL) in 4-hour chromium-release assays (mean killing ± SEM, 2.7 ± 3.1% after incubation of EpCAM positive cell lines with control BiTE®). In contrast, after incubation with solitomab, EpCAM positive USC cells became highly sensitive to T cell cytotoxicity (mean killing ± SEM of 25.7 ± 4.5%; P < 0.0001) by PBL. Ex vivo incubation of autologous tumor associated lymphocytes (TAL) with EpCAM expressing malignant cells in ascites with solitomab, resulted in a significant increase in T-cell proliferation in both CD4+ and CD8+ T cells, increase in T-cell activation markers (i.e., CD25 and HLA-DR), and a reduction in number of viable USC cells in ascites (P < 0.001). CONCLUSIONS: Solitomab induces robust immunologic responses in vitro resulting in increased T-cell activation, proliferation, production of cytokines, and direct killing of tumor cells. These finding suggest that solitomab may represent a novel, potentially effective agent for treatment of recurrent/metastatic and/or chemo-resistant USC overexpressing EpCAM

Foreign policy of the New Left: explaining Brazil's Southern partnerships

Abstract The purpose of this study is to consider the relationship between domestic change and foreign policy in Brazil, a country seeking to become Latin America's hegemon, and achieve greater global status. It focuses on Brazil's partnerships with other countries in the Global South. It argues that, due to the combination of institutions and interests behind foreign policy-making in Brazil, there is no coherent project of South-South engagement. As a result, South-South ties tend to contradict the Brazilian government's foreign policy objective of acting as a global equaliser. The study also examines the drivers of Brazil's foreign conduct, and argues that the politico-economic determinants of foreign policy differ from those of domestic policy