3种国产化试剂盒与Identifiler^TM试剂盒检验结果比较

目的比较3种常用国产化PCR扩增试剂盒与IdentifilerTM试剂盒的检验结果。方法对455份中国汉族无关个体血样,分别用IdentifilerTM、DNA TyperTM15、GoldenEye 16BT、AGCU17＋1 4种试剂盒进行检测,对其中共有的11个基因座位分型结果进行比较,并对有差异的样本进行测序分析。结果 3种国产化试剂盒在455份样本中,发现4例样本的分型结果有差异,差异率为0.88%。其中,DNA TyperTM15试剂盒CSF1PO基因座发现1例等位基因丢失,AGCU17＋1试剂盒D18S51基因座发现1例等位基因扩增不平衡,测序结果表明,2例均系点突变导致;GoldenEye16BT试剂盒D21S11基因座发现2例假3峰型,其中1例测序未发现碱基突变,分析3峰原因可能是stutter滑脱峰所致。结论与IdentifilerTM试剂盒比较,3种国产化试剂盒均有分型差异现象,成因不同,在实践中应给予注意。     

利用16S rDNA序列鉴定常见嗜尸性蝇类种属

目的通过分析16S rDNA 551bp基因序列,鉴定常见嗜尸性蝇类种属。方法随机采集17个地区放置于室外草地的家兔尸体上7个种24只嗜尸性苍蝇样本,经形态学鉴定种类后,提取胸肌DNA,对16S rDNA 551bp基因片段进行PCR扩增,产物纯化、测序后上传GenBank;利用MEGA 4.0软件构建序列间的系统发育树,分析建立种内及种间进化分歧表。结果 24只样本16S rDNA序列分析显示7种蝇类可以较好聚类;其中棕尾别麻蝇种内进化分歧整体均数为2.8%,家蝇为1.5%,丽蝇科的5个种均在0.7%以内。上述7个蝇种的种间进化分歧均数在1.6%～7.1%之间。其中,棕尾别麻蝇、家蝇与其它蝇类的种间分歧均数在4.0%～7.1%之间。结论本文分析结果显示,蝇种间同源性相差明显,采用mtDNA 16S rDNA中551bp基因序列分析,可进行蝇种鉴定。     

Development and validation of the AmpFℓSTR® Identifiler® Direct PCR Amplification Kit: a multiplex assay for the direct amplification of single-source samples

Abstract: The AmpF?STR^® Identifiler^® Direct PCR Amplification Kit is a new short tandem repeat multiplex assay optimized to allow the direct amplification of single‐source blood and buccal samples on FTA^® card without the need for sample purification and quantification. This multiplex assay has been validated according to the FBI/National Standards and SWGDAM guidelines. Validation results revealed that slight variations in primer concentration, master mix component concentration, and thermal cycling parameters did not affect the performance of the chemistry. The assay’s sensitivity was demonstrated by amplifying known amounts of white blood cells spotted onto FTA^® cards, and the assay’s specificity was verified by establishing minimal cross‐reactivity with nonhuman DNA. No effect on the age of the sample stored on the FTA^® substrate was observed and full concordance was established in the population study. These findings of the validation study support the use of the Identifiler^® Direct Kit for forensic standards and database samples genotyping.