高灵敏度薄膜生物传感器基因芯片系统的研制

目的:在传统基因芯片技术基础上,应用生物传感器技术,研制一种能将基因芯片信号原位放大后达肉眼判读灵敏度,无需专用基因芯片检测仪器就可使用,易于在基层医疗单位推广的薄膜生物传感器基因芯片诊断系统.方法:在薄膜生物传感器基片基础上,经过表面化学处理,使特定的基因捕获探针在传感器表面固定,形成特定检测目的生物传感器基因芯片.并以乙肝病毒YMDD区的特异序列设计为例,通过特定的YMDD区的捕获探针,以矩阵的形式点样于传感器芯片表面来实现本系统.同时,纳米金标记的检测探针取代了传统芯片中的荧光标记探针.扩增后的目的PCR片段与捕获探针、生物素标记探针、链亲蛋白纳米金探针进行反应,最后得到探针-生物素-链亲蛋白-纳米金复合物,并在芯片表面经生物传感器芯片将信号原位放大,获得肉眼观的芯片信号并进行分析,完成芯片诊断.以乙型肝炎病毒YMDD突变为例,观察该芯片系统对临床诊断标本诊断的可靠性.结果:基因芯片的检测信号经生物传感器原位放大后能肉眼判读或借助普通数码照像机或计算机扫描,根据信号出现的特定位置即可确定突变的类型.且该生物传感器基因芯片系统信噪比高,在人工合成的寡核苷酸及临床血清的检测中,均可实现生物芯片阴阳性信号完全的有或无的判读;临床血清标本检测证实,使用该传感器芯片系统对前期经过测序确定为YMDD突变的23份临床血清结果与测序结果完全一致.结论:薄膜生物传感器基因芯片集纳米材料、生物传感器技术及原位放大技术为一体,实现了信号的肉眼判读,具有通用性高,准确可靠,无需大型设备,易于在基层医疗单位推广使用.

Chromosomal Localization of Genes bz1,bz2 in Maize by Using Ultra-sensitive FISH with Tyramide Signal Amplification(TSA-FISH)

It has been reported that endosperm undergoes programmed cell death (PCD) during maize kernel development.Both bz1 (bronze ) and bz2 are anthocyanin biosynthetic genes,and related to development of aleuronic layer of maize seeds.Tyramide signal amplification fluorescence in situ hybridization (TSA FISH) is a novel and high sensitive FISH technique,which is suitable for routine application in plant cytogenetic research.Using this technique,we physically mapped the bz1 gene onto the short arm of chromosome 9 and the long arm of chromosome 1;the percentage distances from centromere to hybridization site were 40.2,75.4 respectively,and the bz2 onto the long arm of chromosome 1 and the short arm of chromosome 5;the percentage distances from centromere to hybridization site were 21.6,15.3 separately.The TSA FISH techniques of small low copy DNA sequences for plants are discussed.