Analysis of extrachromosomal homologous recombination in cultured silkworm cells.
Double-strand breaks (DSBs) are potentially lethal lesions causing the loss of chromosomal information. Eukaryotic cells have evolved the error-free repair systems of DSBs by homologous recombination (HR) through gene conversion with or without crossing over. In this study, we have developed a rapid assay system for extrachromosomal HR events in the cultured silkworm BmN4 cells. When HR occurs within the disrupted luciferase gene, an enzymatically active luciferase is restored and expressed. Our results strongly suggest that error-prone single strand annealing (SSA) accounts for the majority of extrachromosomal recombination processes in the cells. However, upon the substrates which cannot be repaired through SSA, DSBs were efficiently repaired though gene conversion. The rapid and sensitive HR assay system developed in the present study is expected to be a powerful tool for the identification and analysis of HR-related genes in the silkworm.
Mon
Hiroaki
H
Laboratory of Silkworm Science, Kyushu University Graduate School of Bioresource and Bioenvironmental Sciences, 6-10-1 Hakozaki, Fukuoka 812-8581, Japan.
Kusakabe
Takahiro
T
Bando
Hisanori
H
Kojima
Katsura
K
Kawaguchi
Yutaka
Y
Koga
Katsumi
K
D000818:Animals; D012831:Bombyx / Q000235:genetics*; D002478:Cells, Cultured; D002875:Chromosomes / Q000235:genetics; D004247:DNA / Q000235:genetics*; D004249:DNA Damage / Q000235:genetics*; D004252:DNA Mutational Analysis / Q000379:methods; D004260:DNA Repair / Q000235:genetics*; D005785:Gene Conversion / Q000235:genetics*; D020869:Gene Expression Profiling / Q000379:methods*; D011995:Recombination, Genetic / Q000235:genetics; D017385:Sequence Homology
D004247:DNA