Mutation of a vitelline membrane protein, BmEP80, is responsible for the silkworm “Ming” lethal egg mutant

• Published in: Gene 2013-02, Vol.515 (2), p.313-319
• Main Authors: Chen, Anli, Gao, Peng, Zhao, Qiaoling, Tang, Shunming, Shen, Xingjia, Zhang, Guozheng, Qiu, Zhiyong, Xia, Dingguo, Huang, Yongping, Xu, Yunmin, He, Ningjia
• Format: Article
• Language: English
• Subjects: 3' Untranslated Regions; Animals; Base Sequence; BmEP80; BmEP80 protein; Bombyx - genetics; Bombyx mori; Chromosome Mapping; chromosomes; DNA; DNA Mutational Analysis; eclosion; Egg Proteins - genetics; Egg Proteins - metabolism; eggs; Female; Gene Expression; genes; Genes, Lethal; Genetic Association Studies; Genetic Linkage; homozygosity; INDEL Mutation; Insect Proteins - genetics; Insect Proteins - metabolism; Lethal egg mutant; Male; membrane proteins; molecular cloning; moths; mutants; mutation; phenotype; polymerase chain reaction; Positional cloning; pupae; RNA; RNA interference; silkworms; two-dimensional gel electrophoresis; vitelline membrane; Vitelline Membrane - metabolism
• ISSN: 0378-1119; 1879-0038
• DOI: 10.1016/j.gene.2012.12.006

The egg stage is an important stage in the silkworm (Bombyx mori) life cycle. Normal silkworm eggs are usually short, elliptical, and laterally flattened, with a sometimes hollowed surface on the lateral side. However, the eggs laid by homozygous recessive “Ming” lethal egg mutants (l-em) lose water and become concaved around 1h, ultimately exhibiting a triangular shape on the egg surfaces. We performed positional cloning, and narrowed down the region containing the gene responsible for the l-em mutant to 360kb on chromosome 10 using 2287 F2 individuals. Using expression analysis and RNA interference, the best l-em candidate gene was shown to be BmEP80. The results of the inverse polymerase chain reaction showed that an ~1.9kb region from the 3′ untranslated region of BmVMP23 to the forepart of BmEP80 was replaced by a >100kb DNA fragment in the l-em mutant. Several eggs laid by the normal moths injected with BmEP80 small interfering RNAs were evidently depressed and exhibited a triangular shape on the surface. The phenotype exhibited was consistent with the eggs laid by the l-em mutant. Moreover, two-dimensional gel electrophoresis showed that the BmEP80 protein was expressed in the ovary from the 9th day of the pupa stage to eclosion in the wild-type silkworm, but was absent in the l-em mutant. These results indicate that BmEP80 is responsible for the l-em mutation. [Display omitted] ► L-em mutant was a new lethal egg mutant of the silkworm, Bombyx mori. ► Mutation of the BmEP80 gene is responsible for the l-em mutant. ► BmEP80 protein is important for maintaining the structural integrity of eggs.