An improved extraction method reveals varied DNA content in different parts of the shells of Pacific oysters

Abstract

The DNA in the shell of Crassostrea gigas could have important roles in the shell biomineralization. However, limited by the low efficiency of existing extraction methods, studies investigating the DNA in shells are lacking. In this study, the shell DNA of C. gigas was extracted using the organic solvent extraction (OSE) and guanidine lysis buffer (GLB) methods; the efficiency and quality of these two methods were compared. The sequences of a mitochondrial gene (cytochrome c oxidase subunit I, COI) and a nuclear gene (28S rRNA) of C. gigas were analyzed to verify the origin of the extracted shell DNA. Finally, the DNA contents of the ventral edge, middle part, and dorsal edge of C. gigas shells were compared. The results showed that OSE had a higher DNA extraction efficiency than GLB; the oyster shell DNA was homologous to the oyster genome; the DNA content was higher in the ventral edge than in the middle part or in the dorsal edge of the C. gigas shell. This study not only reports an improved extraction method for the mollusk shell DNA, but also revealed that the DNA in the oyster shell originates from the oyster body and that the DNA content in different parts of the C. gigas shell showed obvious variance. These results provide supporting evidence for the hypothesis that oyster cells participate in shell formation, and also afford a nondestructive method for oyster genetic identification, which can promote the application of molecular biology technology in oyster breeding. In addition, a shell growth pattern of ‘Under Old & Exceeding Old’ was also proposed.

Advancing responsible genomic analyses of ancient mollusc shells

“…There, geochemical conditions are maintained to allow shell biomineralization by outer mantle epithelial cells beneath the periostracum that serves as a template for shell growth [ 57 ]. In previous work, the periostracum was abraded prior to DNA extraction to remove potential contaminants [ 42 , 45 ], as its high organic content (proteins and polysaccharides) may be prone to microbial colonization, thus reducing the endogenous fraction. However, we observed the opposite here, either because the periostracum is not a good substrate for microbes, or because its chemical compounds prevent bacterial attachment and growth to protect organisms from bacterial fouling [ 87 ].…”

“…X-ray experiments were carried out using the EasyTom XL micro-CT scanner (RX solution) at the Institut de Mécanique des Fluides de Toulouse , France, with a 1.1 mm-wide aluminium filter placed in front of the source. Then, DNA was extracted using the ‘Phenol-Chloroform’ method described in the section ‘Ancient DNA extraction methods’ below, as it was previously shown to efficiently remove co-extracted inhibitors of subsequent enzymatic reactions [ 39 , 42 ].…”

“…Until now, methodological optimization and standards for ethical research have mostly focused on bones and teeth. Meanwhile, mollusc shells have emerged as promising substrates for DNA analyses (e.g., [38][39][40][41][42][43][44][45]) of samples as old as 100,000 years Before Present (yBP; [46]; reviewed in Martin et al [47]). In a similar way to morphological, sclerochronological, sclerochemical, or dating methods commonly applied to ancient mollusc shells from shell middens, refuse dumps, sediment cores or historical collections, aDNA analysis has the potential to provide invaluable evolutionary, ecological and archaeological insights into the impact of climate and environmental changes, as well as into past human activities and resource management strategies: subsistence systems, mobility, migration, exchange networks, aquaculture, production of tools and symbolic artefacts (reviewed in Coutellec [48] and Thomas [49,50]).…”

Anti-Interference Detection of Vibrio parahaemolyticus from Aquatic Food Based on Target-Cyclized RCA with Dynamic Adapter Followed by LAMP

“…Of course, it is possible that some accompanying fragments digested from background DNA are also cyclized by the adapter, but these fragments cannot interfere with the detection due to the following reasons. On the one hand, the probability that fragments from the background DNA have the same two sticky ends as the target is only 1/10 9 , and there is less than one such fragment in every oyster genome (~800 Mb) [28,29]. On the other hand, although these accompanying fragments can be cyclized and pre-amplified by linear RCA, their RCA products cannot be recognized by the LAMP primers and will not influence the specificity of the detection.…”

Molecular phylogeny suggests synonymy of Sandalia bridgesi Lorenz, 2009 with S. triticea (Lamarck, 1810) (Gastropoda, Ovulidae)

“…We have tried but failed to obtain specimens from museums abroad. We also attempted to use the Jiang et al (2019) method to extract DNA from shells of the Institute of Oceanology ( CAS ). As the method requires a minimum of 100 mg of sample and our samples were too small, our attempt failed.…”