国际遗传学杂志
國際遺傳學雜誌
국제유전학잡지
INTERNATIONAL JOURNAL OF GENETICS
2013年
5期
185-190,195
,共7页
张占辉%王小竹%孟岩%闫明%刘卉%韩玲%黄昱%黄尚志%吴白燕
張佔輝%王小竹%孟巖%閆明%劉卉%韓玲%黃昱%黃尚誌%吳白燕
장점휘%왕소죽%맹암%염명%류훼%한령%황욱%황상지%오백연
简并寡核苷酸引物聚合酶链反应%荧光原位杂交%微缺失综合征%嵌合体
簡併寡覈苷痠引物聚閤酶鏈反應%熒光原位雜交%微缺失綜閤徵%嵌閤體
간병과핵감산인물취합매련반응%형광원위잡교%미결실종합정%감합체
Degenerate oligonucleotide primed-PCR (DOP-PCR)%Fluorescence in situhybridization (FISH)%Microdeletion syndrome%Mosaic
目的 优化荧光原位杂交(fluorescence in situ hybridization,FISH)所需探针的制备工作,应用FISH检测染色体7q1 1.2、15q11.2以及22q1 1.2缺失.方法 采用简并寡核苷酸引物聚合酶链式反应(degenerate oligonucleotide primed PCR,DOP-PCR)的方法,扩增人工细菌染色体(bacterial artificial chromosome,BAC) DNA,通过切口平移对扩增的DNA进行间接标记,使用制备好的探针检测染色体微缺失.结果 DOP-PCR方法得到的BAC-DNA扩增产物,经标记后用于FISH,可以观察到清晰且背景低的信号;FISH结果确认了1例染色体7q11.2微缺失嵌合体、1例染色体15q11.2缺失.结论 经过优化的实验方法能够快速制备探针,节约抽提BAC-DNA所需的时间,可以应用于FISH以检测微缺失综合征.
目的 優化熒光原位雜交(fluorescence in situ hybridization,FISH)所需探針的製備工作,應用FISH檢測染色體7q1 1.2、15q11.2以及22q1 1.2缺失.方法 採用簡併寡覈苷痠引物聚閤酶鏈式反應(degenerate oligonucleotide primed PCR,DOP-PCR)的方法,擴增人工細菌染色體(bacterial artificial chromosome,BAC) DNA,通過切口平移對擴增的DNA進行間接標記,使用製備好的探針檢測染色體微缺失.結果 DOP-PCR方法得到的BAC-DNA擴增產物,經標記後用于FISH,可以觀察到清晰且揹景低的信號;FISH結果確認瞭1例染色體7q11.2微缺失嵌閤體、1例染色體15q11.2缺失.結論 經過優化的實驗方法能夠快速製備探針,節約抽提BAC-DNA所需的時間,可以應用于FISH以檢測微缺失綜閤徵.
목적 우화형광원위잡교(fluorescence in situ hybridization,FISH)소수탐침적제비공작,응용FISH검측염색체7q1 1.2、15q11.2이급22q1 1.2결실.방법 채용간병과핵감산인물취합매련식반응(degenerate oligonucleotide primed PCR,DOP-PCR)적방법,확증인공세균염색체(bacterial artificial chromosome,BAC) DNA,통과절구평이대확증적DNA진행간접표기,사용제비호적탐침검측염색체미결실.결과 DOP-PCR방법득도적BAC-DNA확증산물,경표기후용우FISH,가이관찰도청석차배경저적신호;FISH결과학인료1례염색체7q11.2미결실감합체、1례염색체15q11.2결실.결론 경과우화적실험방법능구쾌속제비탐침,절약추제BAC-DNA소수적시간,가이응용우FISH이검측미결실종합정.
Objective To optimize the work of probe preparation used for fluorescence in situ hybridization(FISH) and aplly FISH analysis for the detection of deletions 7q1 1.2,15q11.2,and 22q1 1.2.Methods The DNA of bacterial artificial chromosome (BAC) was amplified by degenerate oligonucleotide primed PCR (DOP-PCR).Indirect labeling of the amplified products was performed by nick translation.Subsequently,the prepared FISH probes were used for detection of chromosome microdeletion.Results By means of DOP-PCR,we got sufficient amounts of FISH probes which provided clear hybridization signals and low backgroud.FISH analysis showed one case with of 15 q 11.2 deletion and another case with a mosaic 7q1 1.2 deletion.Conclusion The optimized method developed by us enables generation of probes to run faster without spending much time on isolating BAC-DNA.The probes could be applied in FISH analysis for the detection of microdeletion syndromes.
