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. 2015 Feb 5;16(1):44.
doi: 10.1186/s12864-015-1241-x.

A de novo transcriptome of the noble scallop, Chlamys nobilis, focusing on mining transcripts for carotenoid-based coloration

Helu Liu  1   2   3 , Huaiping Zheng  4   5 , Hongkuan Zhang  6   7 , Longhui Deng  8   9 , Wenhua Liu  10   11 , Shuqi Wang  12   13 , Fang Meng  14   15 , Yajun Wang  16   17 , Zhicheng Guo  18   19 , Shengkang Li  20   21 , Guofan Zhang  22
Affiliations

A de novo transcriptome of the noble scallop, Chlamys nobilis, focusing on mining transcripts for carotenoid-based coloration

Helu Liu et al. BMC Genomics. .

Abstract

Background: The noble scallop Chlamys nobilis Reeve displays polymorphism in shell and muscle colors. Previous research showed that the orange scallops with orange shell and muscle had a significantly higher carotenoid content than the brown ones with brown shell and white muscle. There is currently a need to identify candidate genes associated with carotenoid-based coloration.

Results: In the present study, 454 GS-FLX sequencing of noble scallop transcriptome yielded 1,181,060 clean sequence reads, which were assembled into 49,717 isotigs, leaving 110,158 reads as the singletons. Of the 159,875 unique sequences, 11.84% isotigs and 9.35% singletons were annotated. Moreover, 3,844 SSRs and over 120,000 high confidence variants (SNPs and INDELs) were identified. Especially, one class B scavenge receptor termed SRB-like-3 was discovered to express only in orange scallops and absent in brown ones, suggesting a significant association with high carotenoid content. Down-regulation of SRB-like-3 mRNA by RNA interference remarkably decreased blood carotenoid, providing compelling evidence that SRB-like-3 is an ideal candidate gene controlling carotenoid deposition and determining orange coloration.

Conclusion: Transcriptome analysis of noble scallop reveals a novel scavenger receptor significantly associated with orange scallop rich in carotenoid content. Our findings pave the way for further functional elucidation of this gene and molecular basis of carotenoid deposition in orange scallop.

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Figures

Figure 1
Figure 1
Line of the C. nobilis was used in transcriptome sequencing. (A) Both parents are orange scallop. (B) Offspring with orange and brown coloration segregation obtained by crossing two orange scallops. (C) Four kinds of tissues: gonad (a), mantle (b), gill (c), and adductor muscle (d). Male scallop has a white or lighter orange color gonad, while female scallop has a heavier orange color gonad.
Figure 2
Figure 2
Overview of the C. nobilis transcriptome sequencing and assembly. (A) Size distribution of 454 raw reads. (B) Size distribution of 454 reads after removal of adaptor and short sequences. (C) Log-log plot showing the dependence of isotigs lengths on the number of reads assembled into each isotigs. (D) Size distribution of isotigs.
Figure 3
Figure 3
Functional annotation of assembled sequences based on gene ontology (GO) categorization. GO analysis was performed at the level 2 for three main categories (cellular component, molecular function and biological process).
Figure 4
Figure 4
Classification of single nucleotide polymorphisms (SNPs) identified from 454 sequences. The overall frequency of these SNP types in C. nobilis transcriptome is one per 278 bp.
Figure 5
Figure 5
Comparison of the expression level of 26 selected tentative carotenoid deposition transcripts in orange and brown scallop adductor muscle. 6 scallops were used in the experiment and each expression analysis was also performed in two independent experiments. Significant difference was performed by one-way ANOVA test (P < 0.05).
Figure 6
Figure 6
Alignment of SRB-like-2 and SRB-like-3. Primers S3F1 and S3R1 give special PCR amplification of SRB-like-3.
Figure 7
Figure 7
Tissue expression profile of SRB-like-3 in orange scallop. Different letter means significant difference by one-way ANOVA test (P < 0.05).
Figure 8
Figure 8
mRNA expression of SRB-like-3 after injection of dsRNA. Letter indicates comparison of the same tissue. Different letter means significant difference by one-way ANOVA test (P < 0.05).
Figure 9
Figure 9
Carotenoids extration from the blood (dsEGFP: dsRNA of EGFP; dsSRB-like-3: dsRNA of SRB-like-3).
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