OBJECTIVETo establish a convenient, practical and high efficient method of DNA extraction of os cervi, and lay the foundation of identification of animal bones.

METHODThe bones of sika deer, red deer, cattle, dog and pig were used to extract DNA under different decalcification time (24,48,72 h) and decalcification temperature (4,25,37,56,70 degrees C), and extract method.

RESULTIt proved by experiments that demineralization process promotes the cracking of osteocyte. In a broad of decalcification time and temperature, DNA could be extracted from all bone samples successfully while the quantity varied slightly.

CONCLUSIONSamples (about 0.1 g) decalcify with 0. mol x L(-1) EDTA at 4 degrees C for 24 h, then water-bath for 1 h after lysis buffer added, DNA extracted via the method above is of high quality and can be used for PCR.

Animals ; Bone Demineralization Technique ; Bone and Bones ; chemistry ; metabolism ; Cattle ; DNA ; isolation & purification ; Deer ; Dogs ; Polymerase Chain Reaction ; Swine ; Temperature ; Time Factors

Objective:To identify the pathogenic mutation in a patient with Oguchi disease.Methods:A Japanese patient with Oguchi disease was enrolled in this study, and underwent a comprehensive medical history assessment and multiple ophthalmic examinations, including BCVA, OCT, color fundus photography and full field electroretinogram. Genomic deoxyribonucleic acid (DNA) was extracted from peripheral blood samples for whole exome sequencing. The gene mutation was detected, and the analysis software was used to determine the conservation of the mutation and the possible structural changes.Results:The patient, 71 years old, with consanguineous parents, complained of night blindness since early childhood. BCVA in both eyes was 0.7 and the golden-yellowish reflex appeared in the grey retina. The scotopic 0.01 ERGs showed a extinguished reaction in both eyes. The scotopic 3.0 ERGs showed a "negative" configuration with a significantly reduced a wave and a nearly absent b wave. A homozygous deletion mutation in the SAG gene (c.924delA, p.N309Tfs*12) in this patient was founded by DNA sequencing, which was predicted to generate prematurely truncated SAG protein and result in severe structural change. Homology analysis of the protein sequence indicated that the mutation resulted in an altered amino acid which was evolutionarily highly conserved among different species, strongly suggesting the potential pathogenicity of this homozygous mutation.Conclusion:The mutation c.924delA(309Tfs*12) in SAG cause Oguchi disease in this patient.