Isolation of sperm vesicles from adult male mayflies and other insects to prepare high molecular weight genomic DNA samples.
We describe here a simple and efficient protocol for genomic DNA isolation from adult males of insects: e.g., Ephemeroptera, Odonata, Orthoptera and Dictyoptera. To minimize contamination of external DNA source, the sperm vesicles were isolated from male individuals from which high molecular weight genomic DNA was extracted. According to this protocol, the genomic DNA samples obtained were high quality (intact), and abundant enough for genotyping analyses and molecular cloning. The protocol reported here enables us to process a huge number of individuals at a time with escaping from cross-contamination, and thus it is quite useful for conducting genetic studies at least in some species of insects. The large yield of high molecular weight DNA from single individual may be advantageous for non PCR-based experiments. As a case study of the protocol, partial coding sequences of histone H3 and EF-1alpha genes are determined for some insects with PCR-amplified DNA fragments.
Takemon
Yasuhiro
Y
Disaster Prevention Research Institute, Kyoto University, Uji, 611-0011, Japan.
Yamamoto
Akiko
A
Nakashima
Masashi
M
Tanida
Kazumi
K
Kishi
Mitsuo
M
Kato
Mikio
M
D000375:Aging / Q000502:physiology; D000818:Animals; D004247:DNA / Q000737:chemistry* / Q000302:isolation & purification*; D049750:Genome, Insect; D007313:Insecta / Q000166:cytology* / Q000235:genetics*; D008297:Male; D008970:Molecular Weight; D010802:Phylogeny; D013094:Spermatozoa / Q000166:cytology*
D004247:DNA