Mentor: Jennifer Schumacher, Ph.D.

Congenital heart defects (CHDs) affect 1% of births, often resulting from dysregulated cardiac development and genetic mutations [1]. We use the transparent zebrafish (Danio rerio) model to analyze orthologus cardiovascular genes that may be associated with CHDs. To accomplish this, we employed co-expression analysis to identify novel regulatory genes, and then analyzed where these genes are expressed. pabpc4 and shdb were identified as candidates due to either co-expression with the ventricular marker vmhc or relation to the she gene family, respiectively. We hypothesized that pabpc4 functions within specific cardiac or striated muscle domains, and that shdb will be expressed as a vascular adaptor protein [3].

Sequences were identified via ZFIN, and primers were designed using Primer3 and ApE. Following PCR amplification of 96 hpf cDNA and subsequent gel purification, purified DNA templates were used to synthesize antisense RNA probes. Gene expression patterns were localized using in situ hybridization (ISH), imaged via Zeiss V8 microscopy, and analyzed qualitatively based on observed staining patterns. pabpc4 was found to be expressed in the fins at 48 hpf, in the yolk at 72 hpf, in the ventrical and the swim bladder at 96 hpf. Expression was found in the somites, facial-cranial muscles and in a peculiar spot on the lateral head of all stages, hypothesized to be a gland. shdb was found to be expressed mainly in the nervous tissue of the zebrafish embryo and particularly in the pectoral fins.

Future research will include investigating the peculiar staining patterns of pabpc4, including more stages in the shdb ISH analysis, and troubleshooting the Proteinase K protocol.

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