Cloning and characterization of a novel gene that is expressed during neural development

• Main Authors: Ting-Bing Wu, 吳庭玢
• Other Authors: Ming-Ji Fann
• Format: Others
• Language: en_US
• Published: 1999
• Online Access: http://ndltd.ncl.edu.tw/handle/31850443808126219012

碩士 === 國立陽明大學 === 神經科學研究所 === 88 === Abstract During early neural development, many intrinsic and extrinsic factors affected the determination of cell fate. A cDNA subtractive hybridization method was adopted to examine genes that are specifically expressed during early neural development in our laboratory. From E10.5 neural tube specific cDNA library we obtained 79 genes which are found no similarity with known genes in the databases at NCBI or the Japan GenomeNet. One clone, E71, was analyzed further. The quantitative RT-PCR demonstrated that the difference of expression of E71 between E10.5 neural tube and P1 brain is 2 folds. To clone the full-length cDNA, the rapid amplification of cDNA ends (RACE) method was performed using several E71-specific primers. The rat E10.5 neural tube total RNA was reverse-transcribed by Superscript II using a E71-specific primer in the presence of a unique 30-base sequence adapter SMARTII. After template-swicthing, the cDNA synthesized by RT can be amplified by PCR using the adapter and E71-specific primers. A clone with 3842 bp insert was obtained after 3 rounds of RACE. An open reading frame between 325 bp to 672 bp was found. By Northern blot, I found that E71 has two transcripts. One is about 3.6 kb and the other is about 1.3 kb. E71 is expressed ubiquitously in various organs and in the CNS. Further analysis is needed to characterize function of E71.