Study And Analysis On Expressed Sequenced Tags From Chicken Hypothalamus And Pig Fat Tissue

The generation of expressed sequence tags (EST5) has been proven to be a rapid and efficient approach by which to identify expressed genes and characterize the information content of genome. To this aim, we constructed a eDNA library and generated 83 ESTs from chicken hypothalamus tissue. The titer of our library constructed was 3.8x 106, and the insert fragment length was 1.6kb. All the 83 ESTs generated from chicken hypothalamus have been subscribed to genbank and gotten the Acce. No.. Of these ESTs, 38 matched to function-known genes by blastn in genbank non-redundant nucleotide database; 12 matched only to other known ESTs by blastn in EST database; 21 showed no match to any ESTs or function-known genes and others were mitochondrial genes. This research also found several nervous system specific genes such as tyrosine 3-monooxygenase by expression profile construction. The homologous analysis with protein database by tblastx showed that there were many homologous ORF multialigned with known and unknown ESTs sequenced. In the result, 7790 high quality reads were obtained from porcine fat tissue cDNA library by large scale sequencing. Of these ESTs, 4461 matched to function-known genes; 1000 matched only to other known ESTs; 1033 showed no match to any ESTs or function-known genes; and others were mitochondrial genes or pollution sequences. The construction of expression profiles of function-known genes made us find 339 clones and 92 lipid-metabolism-related genes including many tissue specific multicopy genes expressed in porcine fat tissue such as adipocyte fatty acid binding protein (aP2, 48 copies). All these genes could be studied as candidate genes controlling the pig fat traits. The homology analysis with protein database showed that there were many homologous ORF multialigned with known and unknown ESTs. The contig assembling results with different standards showed that the phrap software popularly used in genome analysis didn't suit for EST analysis of this test. By combining Accession No. index to cluster function-known genes with CAT software to cluster other ESTs, a new method was used to identify 752 unique clusters comprising 3259 ESTs, whereas the remaining 3235 ESTs were singlets, and the estimated number of unique genes identified in the porcine fat ESTs data set was approximately 3987. dbSTS blastn in STS database results showed that 592 and 44 matched to known STSs in function-known ESTs and known ESTs, respectively, and distributed on many porcine chromosomes including chromosome 1, 2, 3, 6, 7, 9, 10, 12, 13, 14, 15, 17, 18, which can be used to pig map construction and comparative mapping analysis. With the constructed linking sequence (l0,340bp) with ESTs of 70 function-known genes, we analyzed the genetic relationships among four mammalian species and the results showed that the nearest genetic relationship with pig is bovine, the next is human, and the farthest is mouse. The analysis also showed that the cDNA/ESTs sequence can also be used to make evolutional analysis as genome methods. The analysis of SNPs from ESTs made us identify 391 SNPs and most of them were CiT and A/G transition with a higher frequency; and it also showed that mining SNPs from ESTs is a rapid and efficient method to find new SNPs from genome.