Specific Polymerase Chain Reaction Molecular Identification of Pheretima aspergillum (E. Perrier) / 广州中医药大学学报

Objective To screen out the polymerase chain reaction ( PCR) primers for specifically identifying Pheretima aspergillum (E. Perrier) , so as to establish a rapid and accurate method to identify the origin of Pheretima aspergillum. Methods Four kinds of earthworms recorded in pharmacopoeia and six kinds of their adulterants commonly seen in the market were collected. The 12SrRNA sequences related to earthworm were downloaded from GenBank database. PCR amplification for ten kinds of samples was performed using the universal primers, and then the products were sequenced. According to the differences between the above sequences, three pairs of specific primers in the non-conservative district were designed for identification of Pheretima aspergillum. Results High-specificity PCR amplification for Pheretima aspergillum with 12St/12Stf primer occurred when the annealing temperature was increased to 64℃, and only Pheretima aspergillum had single amplification strip, while no strips were found in the other adulterants under the same condition. The achieved specific primers could be well verified in 15 kinds of medicinal materials of earthworm purchased in the market, which had the same morphological features showed by the traditional identifying method. Conclusion With 12St/12Stf primer as a specific marker, the identifica tion of Pheretima aspergillum is rapid and accurate in related species of Pheretima aspergillum, and avoids the effect of some factors such as integrity of experimental materials and drying process.

Prevalence Study of GJB2,SLC26A4 and mtDNA 12S rRNA1555A>G Mutations in Hui Ethnic Group Patients with Non-Syndromic Hearing Loss from Northwest China / 听力学及言语疾病杂志

Objective To investigate the prevalence of GJB2 ,SLC26A4 and mitochondrial DNA 12S rRNA m .1555A>G(mtDNA 1555A>G) mutations in Hui ethic group patients with nonsyndromic hearing loss (NSHL) from Northwest China .Methods A total of 420 peripheral blood samples were collected from unrelated Hui ethic group probands with NSHL in Northwest China .Amplified the target gene by polymerase chain reaction (PCR) af-ter extracting genomic DNA from whole blood .The mtDNA 1555A>G mutation was detected by PCR -Alw26I di-gestion ,then direct sequencing was used to the positive samples of mtDNA 1555A> G ,the coding region of GJB2 gene ,exon 8 and 19 of SLC26A4 gene .Results There were 11(2 .62% ) cases caused by mtDNA 1555A>G homo-zygous mutation in 420 patients with NSHL .There were 41(9 .76% ) cases including homozygote and compound het-erozygote ,caused by GJB2 gene mutation ,which was the most frequent deafness -related gene .The allel frequency of c .235delC accounted for 6 .90% ,as well as the most frequent(51 .33% ) mutational pattern in GJB2 gene .There were 20 patients(4 .76% ) were found carring two allel mutations in SLC26A4 gene .The allel frequency of c .919 -2A>G was 5 .0% ,accounting for a total of 68 .85% in all base alterations of SLC26A4 gene ,which was the major mutant form of SLC26A4 gene .Conclusion GJB2 gene is the most common deafness -gene in Hui ethnic group pa-tients with NSHL from Northwest China ,while c .235delC is the main mutant form ,and c .919-2A>G is the hot-spot mutation of SLC26A4 gene .Through this study we can provide the molecular epidemiology basis for Hui ethnic group patients with NSHL from Northwest China in genetic diagnosis ,genetic counseling and therapy by associated testing of three frequent hearing loss genes .