0125, i.e.: 0.05/4 markers. We documented the development of gross edema at 4 and free copy 6 days post-fertilization in live embryos. We performed dextran clearance experiments following previously described protocols [55]. Briefly, 80 hours after MO injection, we anesthetized embryos in 4 mg/ml Tricaine in embryo water (120 dilution), then positioned embryos on their back in a 1% agarose injection mold. We injected an equal volume of tetramethylrhodamine dextran (70,000 MW; Invitrogen) into the cardiac sinus venosus of each embryo. We then returned the embryos to fresh embryo water. Using fluorescence microscopy, we imaged the embryos at 2 hours post-injection (82 hpf) to demonstrate equal loading, then at 48 hours post-injection (128 hpf) to evaluate dextran clearance.

Embryos were injected with control, mpped2, or casp9 MOs at the one-cell stage. At 48 hpf, embryos were manually dechorionated, anesthetized in a 120 dilution of 4 mg/ml Tricaine in embryo water, and oriented on a 1% agarose injection mold. As previously described [56], embryos were injected with equal volumes of 10 mg/ml gentamicin (Sigma) in the cardiac sinus venosus, returned to fresh embryo water, and subsequently scored for edema (prevalence, time of onset) over the next 3 days. Supporting Information Figure S1 Flowchart of the project. (TIF) Click here for additional data file.(256K, tif) Figure S2 Genome-wide ?log10 P values plot from stage 1 discovery meta-analysis. Plots show the discovery analysis of eGFRcrea in the overall group, with known loci [8], [9] highlighted in orange and novel loci highlighted in blue (A), and in strata of the main CKD risk factors (B, C, D, and E), with complementary groups being contrasted each other.

The dotted line indicates the genome-wide significance threshold at P value=5��10?8. The unmarked locus is RNASEH2C on chromosome 11, colored in gray despite genome-wide significance. The P value for the current stage 1 discovery for rs4014195 was 2.7��10?9. This locus previously did not replicate [9]; when we additionally considered our prior non-overlapping in silico and de novo replication Cilengitide data, the current stage 2 P value was 0.8832, yielding a combined stage 1+stage 2 P value of 2.6��10?7. Therefore, we did not submit this SNP for further replication. (PDF) Click here for additional data file.(724K, pdf) Figure S3 Quantile-quantile plots of observed versus expected ?log10 P values from the discovery analysis of eGFRcrea overall (A) and by strata of the main CKD risk factors (B).