Dissertation > Excellent graduate degree dissertation topics show

Identification of a Novel Exon in the Disease Gene FMR1 of Fragile X Syndrome

Author: LiaoJuan
Tutor: LanFengHua
School: Fujian Medical
Course: Clinical Laboratory Science
Keywords: FMR1 gene Alternative splicing Bioinformatics Exon Semi-nested PCR
CLC: R596.1
Type: Master's thesis
Year: 2011
Downloads: 35
Quote: 0
Read: Download Dissertation

Abstract

Fragile X syndrome (fragile X syndrome, FXS) An incidence of mental retardation after Down's syndrome, their causative gene FMR1 (fragile X mental retardation 1 gene) is located on chromosome X27.3 17 exons and 16 introns, spanning 38kB. Has confirmed that there are multiple splicing FMR1 gene, mainly involving 12,14,15,17 four exons spliced ??forms and in different tissues is not consistent, and various splicing isoforms (splice isoform), the specific function is not clear . Due to the higher incidence of FXS also complex and diverse clinical manifestations, the analysis of the time and space characteristics of alternative splicing of the FMR1 gene expression is a new starting point for the pathogenesis study the FMRP protein function and FXS. Before found in a suspected the FXS patient's FMR1 gene in the laboratory for a 46bp size of the new occult exon (cryptic exon), between exon 9 and exon 10 intron 9 small fragments. The study occult exon also exists in other species, we take advantage of bioinformatics software FMR1 gene analysis of other species. The results showed that: a similar gene sequences homologous genes in the rhesus monkey, chimpanzee, Sumatran orangutan, but their position is different, and no similar mouse and rat genome homologous sequences. Splice variants of the human FMR1 gene may predict. On this basis, the basis for a new type of occult outer exon characteristics, using semi-nested PCR, detection of this new type of occult exon cDNA samples of 27 healthy subjects and five kinds of human cDNA library from the RNA level, the results show : 27 parts normal cDNA samples, and five kinds of a human cDNA library, there are new occult exon sequences. In this study, the use of hybrid minigene splicing assay, the new suspected FXS patients and a normal human peripheral blood FMR1 gene occult exon expression analysis from the cellular level. Using PCR amplification of the FMR1 gene intron 9 full-length and its adjacent exon 9 and exon 10 partial sequence inserted into the pcDNA ^{TM 3.1/Hygro () vector and transiently transfected HeLa cells 48 hours after the extraction of HeLa cell total RNA by RT-PCR analysis of this new alternative splicing product expression or not. The results: patients with normal samples and the recombinant vector insert gene FMR1 gene fragment sequencing results with GenBank sequences, but three sites base change in the patient samples constructed recombinant vector inserted fragment sequencing results : 21452 base C → T the 21513 nucleotide C → T, T → C 23784 base change, the first two nucleotide sites were polymorphic, and 23,784 base T → C change , from bioinformatics software analysis results, may ISE, resulting 46bp sequence in the splicing process is preserved, and normal human samples constructed recombinant vector inserted fragment sequencing results, two loci alkali base changes, i.e.: 21452 base C → T, 21513 nucleotide C → T. Nevertheless, the semi-nested PCR product sequencing, the recombinant vector of the patients and normal source of transcripts in HeLa cells were detected in the occult exon. This study demonstrates that, variable between splicing with FXS laid the foundation for the study of the FMR1 gene intron 9 in the presence of a new conceal alternatively spliced ??exons in the human FMR1 gene.}

Related Dissertations

CLC: > Medicine, health > Internal Medicine > Systemic disease > Genetic diseases > Chromosomal diseases
© 2012 www.DissertationTopic.Net Mobile