阿尔茨海默症相关基因的克隆和细胞模型的建立.pdfVIP

?????????? ???λ???? ?????????????????????????????? ???????????? ?????λ?????? ??????????????????? ?????????????;???????????? ?????????????(Alzheimer??S disease??AD)???????????????????? ????????????????????о????????ж????????????????????????з???? ????????????????????????????????о????????????????????? ?????????????????????????á????о????RT??PCR???????????????? ???????AD????????????????pcDNA3??1????????徭??????? U251?????G418????????APPsw(Lys595Asn??Met596Leu)???????BACEl ?????????????????????DNA PCR??RT??PCR??Western blot???????? ??????????????mRNA???????????????APPsw?????BACEl???? ????U251???????????????APPsw(Lys595Asn??Met596Leu)????APP?? ???Abeta????????????????????????BACEl?????Abeta?????????? ?????о???????????????о????????????????????????????? ????????? ????????????????????????APP??BACEl ??????Q421 ABSTRACT ABSTRACT??Alzheimegs disease(AD)is the most frequent neurodegenerative disorder in the aged population??which Call reduce the memory capability,sensibility capability, judgement capability and motor ability??Establishing a cell model of Alzheimers disease is very useful for studying the mechanism of AD pathogenesis and AD pathologic process??and it is also very critical for screening new drugs which Can cure AD??In this research we cloned ten kinds of genes associated with AD pathogenesis by RT??PCR from human neuron??With the help of lipofectamine 2000??cultured U25 1 cells were transfected by pcDNA3??1 containing mutant APPsw(Lys595Asn??Met596Leu)and wild BACElgene separately??The positive cells were screened out by G418??The results of PCR of genomic DNA??RT-PCR and Western blowing showed that the U251 cells transfected by APPsw and BACE 1 Can stably express the relevant mRNA and protein?? However,there is no obviously effect observed on cell morphology??In the U251 cells transfected by APPsw??the expressing level of the APP and resulted Abeta age lfled in contrast to control clones??Similarly,in the U251 cells transfected by BACE1??the product of the