Validating real-time PCR and field manipulations using the coral Montipora Capitata

Abstract

In this study, qRT -PCR amplification of the gene encoding hsp70 from Montipora capitata and Symbiodinium were tested and normalized using the SMP method (Mayfield et al. accepted). The goal of this work was to confirm the utility of the exogenous spike as a reference gene for calculating SMP and as a housekeeping gene in qRT-PCR amplification of cDNA as well as to validate the general methods utilized for qRT-PCR using corals (the RNA extraction, cDNA synthesis, and amplification steps). Two field experiments were also designed to explore how the Symbiodinium residing in coral hosts responded to 1) fragmentation and recuperation and 2) transplantation by tracking symbiont photophysiological data and changes in Symbiodinium cell densities using qRT-PCR. Temperature and light intensity data collected from the transplantation experiment were used to determine whether control depth and experimental depth were statistically different. Coral fragments used in the field experiments were also used to test qRT-PCR methods.