The Consrtuction Of Four STR Locus Allelic Ladder By Using The Cloning Technique And Forensic Evalution

Objective:To resolve the problem of the accuracy and standardization of STR-PCR typing in forensic practice .we choose D5S818, D19S253, DYS447,DYS448 four STR locus and explore a method to make standard allelic ladder by using the cloning technique .The D19S253 allilic ladder we produced have been used in genetic polymorphism study and forensic case evalution.Method: The DNA were extracted with a reform method of chloroform phenol . STR-PCR amplification,analysis is using non-denaturing polyacylaminde gel electrophoresis and sliver staining .Cloning technique and ABI377 sequencer were carried out from separating four locus alleles of 487 individuals.The different allelic fragments were isolated from the gel ,eluted into the distilled water and re-amplified by PCR. The purified allelic fragment were connected with PGEM-T vectors directly. The combinants were transfected into the component E.coil Top10' cells follwed by cloning and plasmid purification. The four locus allelic ladders were produced by re-amplifying the recombinant plasmid DNA.ResuIts:The four locus (D5S818,D19S253,DYS447,DYS448 )allelic ladder made in this way can be produced in a large amount and stored in a relatively long period .Conclusion:The result demonstrated that the four locus standard allelic ladder produced by the cloning technique are practical in forensic science application.