Mutation in SRBI gene and its association with the red shell color in Meretrix meretrix

The hard clam (Meretrix meretrix) is one of the most commercially important cultured shellfish in China. The red shell clam is richer in carotenoids, which makes its shell color red and has higher nutritional value compared with the white shell clam. In order to understand the mutation of scavenger receptor class B type I (SRBI) and its association with the formation of red shell color in M. meretrix, we analyzed the SNPs of Mm-SRBI gene CDS by using direct sequencing among 181 red shell clams and 207 white shell clams. Immunofluorescence (IF) and Western blot (WB) were used to analyze the expression characteristics of Mm-SRBI protein in the mantle of two shell color strains. The results showed that a total of 15 single nucleotide polymorphisms (SNPs) were detected, and there were only two types of mutation in Mm-SRBI: transition and transversion, and the ratio of the two was about 3∶1. Among the 15 loci, 4 loci were significantly associated with shell color. The statistical results indicated that in red shell strain the range of H_o was 0.309 4−0.442 0, H_e 0.468 8−0.494 6, and N_e 1.863 4−1.973 4, while in the white shell strain H_o was 0.231 9−0.328 5, H_e 0.324 4−0.500 0, and N_e 1.639 9−1.995 3. The PIC value evidenced that these loci were all moderately polymorphic (0.5>PIC>0.25). Additionally, all of them were in conserved sequences, in which c.723 A/G site was non-synonymous mutation that led to amino acid changes Ile723Val, and 4 loci were genetically strongly linked (D′>0.75). The results of IF and WB revealed that Mm-SRBI protein was higher expressed in the mantle of red-shelled clams compared to that of the white-shelled clams and was mainly expressed at the outer epithelium of the mantle. Moreover, the quantitative analysis of gray values demonstrated that Mm-SRBI protein expression in red-shell clams was approximately 4.5 times higher than that in white-shell clams. In summary, our results suggested that the mutations of Mm-SRBI and the high expression of its protein may lead to differences in the metabolism of carotenoids in M. meretrix, which may further lead to the formation of red shell color. It also provided a helpful basis to explore the molecular mechanisms of carotenoids metabolism underlying shell coloration and could be potentially applied to marker-assisted selection breeding programs for M. meretrix.