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The frozen T. maxima mantle tissue samples were homogenized using a Freezer/Mill (Model 6875, SPEX Sample Prep, USA) by grounding up the tissues in liquid nitrogen at a rate of ten impacts per second for a total of 90 seconds. The ground tissues were then dissolved in 5 ml MilliQ water (sterilized under UV light for one hour) and the homogenate was transferred to 1.5 ml Eppendorf tubes. Samples were frozen at -20 C until further processing. DNA extractions from the tissue samples were extracted with the Qiagen DNeasy Blood & Tissue Kit (Qiagen, Hilden, Germany) following the manufacturer's instructions with minor modifications. Briefly, 90 l of the tissue homogenate was added to 1.5 ml Eppendorf tubes containing 90 l of ATL buffer and 20 l of Proteinase K and incubated at 56C for 1 hr. DNA extractions were then continued according to manufacturers instructions.DNA concentrations were measured using a Qubit 2.0 fluorometer (ThermoFisher Scientific), and samples were adjusted to 10 ng/l. PCRs were conducted using the SYM_VAR_5.8S2 and SYM_VAR_REV primer pair. For all samples, triplicate PCRs were performed using 10 ng of DNA using the Qiagen Multiplex PCR kit and a final primer concentration of 0.3 M in a reaction volume of 10 l. Thermal cycling conditions were: 95C for 15 min, followed by 27 cycles of 95C for 30 s, 55C for 90 s, 72C for 30 s, and a final extension cycle of 72C at 10 min. Five L of the PCR were run on a 1% agarose gel to visualize amplification.