Genetic Quality Detection Of Inbred Mice DNA

There are thousands of mice stains such as closed strain,inbred strain,congenic strain,recombinant inbred strain and mutant strain and so on since laboratory mice have been a research tool.The mutant speed of laboratory mice is fast,and the mutations do not show in the characters.With the development of biomedicine,either in academic or in industry,the quality of laboratory animals is becoming more and more important,and the genetic monitoring methods of laboratory animal have higher requirements.At present,the genetic monitoring in the standards related to genetic quality control has some limitations and it is inadequate in the comprehensive reflection of experimental animal genetic profile.In addition,the program of experimental animal genetic quality which International Association of experimental animals suggested is DNA detection(http://www.iclas.org/).Using DNA markers detection technique to determine of the genetic background of mice strains is current trend.In 2002,International Mouse Genome Sequencing Consortium completed the total sequencing of C57BL/6J(classic laboratory strain,B6).The same year,Celera also completed the sequence draft of another three inbred strains.Subsequently,more and more mice strains have been heavy sequencing.The results show that 2/3 regions of the laboratory mouse genome has low density SNP(0.5/10kb),and the high density SNP(40/10kb)covers the rest of the 1/3 region.Because of the SNP number is great and the cost of high-throughput SNP genotyping is low,the world's leading experimental animal companies have already used SNP for genetic testing.Methods Four groups of multiple PCR-LDR(Polymerase chain reaction and ligase detection reaction,PCR-LDR)genotyping panels were constructed for common inbred mice in this article.45 SNP from 21 chromosomes were selected as targets,as well as primers and probes were designed specific to these SNP,respectively.First,the small amount of genomic DNA rapid extraction kit for DNA extraction in inbred mouse.Second,a preliminary 2%agarose gel electrophoresis for detecting the products of foursets multiplex PCR amplification.Then,the standard 6%denaturing sequencing PAGE gel electrophoresis for LDR products.Last,GeneScanTM 672 for collecting data and Genemapper for genotyping.Results Annealing temperatures of multiplex PCR amplification of four panels were optimized at 56?.The results showed that 34 SNPs are consistent with the information of the NCBI;2 SNPs have no information;11 SNP is different by sequencing;and 1 SNP is found heterozygous between BALB/c and FVB,it is actually homozygous by sequencing.In this study,three batches of inbred mice samples(66)were tested,the results showed that SNP information of the same strain from different sources is consistent.The probability of appearing 47 SNP information was 13.6%,the probability of appearing 46 SNP information was 36.4%,the probability of appearing 45 SNP information was 90.9%and the probability of appearing 44 SNP information is 100%.In the 10 inbred mice,the biggest difference loci number between every strain pair is 29,the smallest is 7 and the mean is 19.The first three groups can do the identification independently in the inbred strains,the fourth group can do the identification independently in the inbred stains besides C3H and CBA.This showed that the 48 SNP in these mice strains is efficient.In conclusion,our results indicate that PCR-LDR could be used to detect the genetic quality of inbred mice strains,with convenience and high throughout.