False negative rates in Drosophila cell-based RNAi screens: a case study

<p>Abstract</p> <p>Background</p> <p>High-throughput screening using RNAi is a powerful gene discovery method but is often complicated by false positive and false negative results. Whereas false positive results associated with RNAi reagents has been a matter of extensive study, the issue of false negatives has received less attention.</p> <p>Results</p> <p>We performed a meta-analysis of several genome-wide, cell-based <it>Drosophila </it>RNAi screens, together with a more focused RNAi screen, and conclude that the rate of false negative results is at least 8%. Further, we demonstrate how knowledge of the cell transcriptome can be used to resolve ambiguous results and how the number of false negative results can be reduced by using multiple, independently-tested RNAi reagents per gene.</p> <p>Conclusions</p> <p>RNAi reagents that target the same gene do not always yield consistent results due to false positives and weak or ineffective reagents. False positive results can be partially minimized by filtering with transcriptome data. RNAi libraries with multiple reagents per gene also reduce false positive and false negative outcomes when inconsistent results are disambiguated carefully.</p

Mutations in a gene encoding an ABC transporter cause pseudoxanthoma elasticum

Pseudoxanthoma elasticum (PXE) is a heritable disorder characterized by calcification of elastic fibres in skin, arteries and retina that results in dermal lesions with associated laxity and loss of elasticity, arterial insufficiency and retinal haemorrhages leading to macular degefieration(1-5). PXE is usually found as a sporadic disorder, but examples of both autosomal recessive and autosomal dominant forms of PXE have been observed(6). Partial manifestations of the PXE phenotype have also been described in presumed carriers in PXE families(7,8). Linkage of both dominant and recessive forms of PXE to a 5-cM domain on chromosome 16013.1 has been reported (refs 8,9). We have refined this locus to an 820-kb region containing 6 candidate genes(10). Here we report the exclusion of five of these genes and the identification of the first mutations responsible for the development of PXE in a gene encoding a protein associated with multidrug resistance (ABCC6)

CXCR4, CXCR5 and CD44 May Be Involved in Homing of Lymphoma Cells into the Eye in a Patient Derived Xenograft Homing Mouse Model for Primary Vitreoretinal Lymphoma

Background: Primary vitreoretinal lymphoma (PVRL), a rare malignancy of the eye, is strongly related to primary central nervous system lymphoma (PCNSL). We hypothesized that lymphoma cells disseminate to the CNS and eye tissue via distinct homing receptors. The objective of this study was to test expression of CXCR4, CXCR5, CXCR7 and CD44 homing receptors on CD20 positive B-lymphoma cells on enucleated eyes using a PCNSL xenograft mouse model. Methods: We used indirect immunofluorescence double staining for CD20/CXCR4, CD20/CXCR5, CD20/CXCR7 and CD20/CD44 on enucleated eyes of a PCNSL xenograft mouse model with PVRL phenotype (PCNSL group) in comparison to a secondary CNS lymphoma xenograft mouse model (SCNSL group). Lymphoma infiltration was evaluated with an immunoreactive score (IRS). Results: 11/13 paired eyes of the PCNSL but none of the SCNSL group were infiltrated by CD20-positive cells. Particularly the choroid and to a lesser extent the retina of the PCNSL group were infiltrated by CD20+/CXCR4+, CD20+/CXCR5+, few CD20+/CD44+ but no CD20+/CXCR7+ cells. Expression of CXCR4 (p = 0.0205), CXCR5 (p = 0.0004) and CD44 (p &lt; 0.0001) was significantly increased in the PCNSL compared to the SCNSL group. Conclusions: CD20+ PCNSL lymphoma cells infiltrating the eye co-express distinct homing receptors such as CXCR4 and CXCR5 in a PVRL homing mouse model. These receptors may be involved in PVRL homing into the eye

Three-Dimensional Momentum Imaging of Electron Wave Packet Interference in Few-Cycle Laser Pulses

Using a reaction microscope, three-dimensional (3D) electron (and ion) momentum (P) spectra have been recorded for carrier-envelope-phase (CEP) stabilized few-cycle (~5 fs), intense (~4×1014 W/cm2) laser pulses (740 nm) impinging on He. Preferential emission of low-energy electrons (Ee<15 eV) to either hemisphere is observed as a function of the CEP. Clear interference patterns emerge in P space at CEPs with maximum asymmetry, interpreted as attosecond interferences of rescattered and directly emitted electron wave packets by means of a simple model