An in silico approach was developed to survey the genes expressed in four internal organs of pig: liver, kidney, spleen and small intestine. The major procedures of the approach included: (1) BLAST searching against GenBank "est_others" database using human cDNA sequences as queries to screen the po...
An in silico approach was developed to survey the genes expressed in four internal organs of pig: liver, kidney, spleen and small intestine. The major procedures of the approach included: (1) BLAST searching against GenBank "est_others" database using human cDNA sequences as queries to screen the porcine orthologous expressed sequence tags (ESTs), (2) classifying the porcine ESTs records by resources according to certain criteria and (3) analyzing data for ESTs specifically expressed in each organ. In order to do so, four Java programs were developed. Based on the ESTs available in the GenBank database, it was found that there were at least 2,100 genes expressed in these four organs, including 128 in the liver, 81 in the kidney, 780 in the spleen, and 1,423 in the small intestine respectively (a few genes co-expressed in these tissues). Gene expression patterns, such as co-expressed genes, preferentially expressed genes and basic active genes were also compared and characterized among these organs. This study provides a comprehensive model on how to use the bioinformatics approach and Genbank databases to facilitate the discovery of new genes in livestock species.
An in silico approach was developed to survey the genes expressed in four internal organs of pig: liver, kidney, spleen and small intestine. The major procedures of the approach included: (1) BLAST searching against GenBank "est_others" database using human cDNA sequences as queries to screen the porcine orthologous expressed sequence tags (ESTs), (2) classifying the porcine ESTs records by resources according to certain criteria and (3) analyzing data for ESTs specifically expressed in each organ. In order to do so, four Java programs were developed. Based on the ESTs available in the GenBank database, it was found that there were at least 2,100 genes expressed in these four organs, including 128 in the liver, 81 in the kidney, 780 in the spleen, and 1,423 in the small intestine respectively (a few genes co-expressed in these tissues). Gene expression patterns, such as co-expressed genes, preferentially expressed genes and basic active genes were also compared and characterized among these organs. This study provides a comprehensive model on how to use the bioinformatics approach and Genbank databases to facilitate the discovery of new genes in livestock species.
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제안 방법
(1) counting the human genes and their homologous porcine ESTs expressed in the liver, kidney, spleen, and small intestine according to the human chromosome number, and identifying the extremely significant match records in the respective tissues (represented by the top 10).
In this study, we built up an in silica approach to identify the homologies between cDNA sequences of human genes and porcine ESTs sequences by utilizing the present software tools and developing Java programs. First, a standalone BLAST searching program was installed and used to perform BLAST searches to annotate porcine ESTs expressed in the major internal organs. The major procedures included firstly: (1) BLAST searching against GenBank “est_others” database using human cDNA sequences as queries, (2) identifying and screening the porcine orthologous ESTs, (3) classifying the porcine ESTs records by resources according to certain criteria, (4) collecting and arranging data for ESTs specifically expressed in the major internal organs.
Realizing these, we developed an in silico study here. In this study, retrieving homologous sequences between human genes and porcine ESTs sequences by BLAST, allowed us to annotate the porcine ESTs derived from the major internal organs by a fresh gene oriented approach which characterized genes expressed in some tissues from genes to ESTs. This approach provided tools and information for rapid discovery of genes of interest.
In this study, we built up an in silica approach to identify the homologies between cDNA sequences of human genes and porcine ESTs sequences by utilizing the present software tools and developing Java programs. First, a standalone BLAST searching program was installed and used to perform BLAST searches to annotate porcine ESTs expressed in the major internal organs.
The major procedures included firstly: (1) BLAST searching against GenBank “est_others” database using human cDNA sequences as queries, (2) identifying and screening the porcine orthologous ESTs, (3) classifying the porcine ESTs records by resources according to certain criteria, (4) collecting and arranging data for ESTs specifically expressed in the major internal organs. Secondly, four Java programs were developed for sequences collection, sequences alignment and data processing, which included: (1) Web robot, used for collecting mRNA accession numbers by detecting all human chromosome information web pages, (2) Light query, used for automatically querying the NCBI search engine (http://wwww.ncbi.nlm.nih.gov/entrez/) for the FASTA sequences using mRNA accession numbers as search fields, (3) BlastFilter, used to filter out the BLAST matches that do not represent the porcine ESTs or do not meet the requirements with sequence identity by higher than 80% within a continuous alignment of sequences longer than 100 bp, (4) Parser/Screen, used to process Plain text in BLAST research result pages so as to analyze more conveniently and easily, and ultimately form the Microsoft Excel format tables.
Secondly, four Java programs were developed for sequences collection, sequences alignment and data processing, which included: (1) Web robot, used for collecting mRNA accession numbers by detecting all human chromosome information web pages, (2) Light query, used for automatically querying the NCBI search engine (http://wwww.ncbi.nlm.nih.gov/entrez/) for the FASTA sequences using mRNA accession numbers as search fields, (3) BlastFilter, used to filter out the BLAST matches that do not represent the porcine ESTs or do not meet the requirements with sequence identity by higher than 80% within a continuous alignment of sequences longer than 100 bp, (4) Parser/Screen, used to process Plain text in BLAST research result pages so as to analyze more conveniently and easily, and ultimately form the Microsoft Excel format tables.
So for database miners or users, there are three challenges to confront: (1) making full and efficient use of the present software tools resources on-line or off-line in sequence analysis, genetic analysis, and data processing etc. (2) developing a certain program for large-scale in silico study, e.g., four programs 'Web robot’, 'Light query’, 'BlastFilter’, and 'Parser/Screen’ were developed by Java in this study.
To detect the gene expression profiles in the major internal organs of the pig, we have developed an in silico approach for mining the NCBI (The National Center for Biotechnology Information) porcine EST sequence resources using the human genome cDNA sequences as references. The information reported in this paper should be useful for researchers in the field to analyze genes and proteins of their own interest, and to study comparative and functional genomics.
The major procedures included firstly: (1) BLAST searching against GenBank “est_others” database using human cDNA sequences as queries, (2) identifying and screening the porcine orthologous ESTs, (3) classifying the porcine ESTs records by resources according to certain criteria, (4) collecting and arranging data for ESTs specifically expressed in the major internal organs.
First, a standalone BLAST searching program was installed and used to perform BLAST searches to annotate porcine ESTs expressed in the major internal organs. The major procedures included firstly: (1) BLAST searching against GenBank “est_others” database using human cDNA sequences as queries, (2) identifying and screening the porcine orthologous ESTs, (3) classifying the porcine ESTs records by resources according to certain criteria, (4) collecting and arranging data for ESTs specifically expressed in the major internal organs. Secondly, four Java programs were developed for sequences collection, sequences alignment and data processing, which included: (1) Web robot, used for collecting mRNA accession numbers by detecting all human chromosome information web pages, (2) Light query, used for automatically querying the NCBI search engine (http://wwww.
성능/효과
(3) investigating the basic active genes (BAGs) expressed in the major internal organs of pig by setting certain criteria (records were arranged by the number of the porcine EST hit(s) per human gene, and then the cut-off criteria to choose the basic active genes were selected at 3 EST hits (about 0.1% of their total hits) per gene.
Among them, the difference between liver and kidney reached the 0.05 significance level (p<0.05, t=2.806), and all the remainder reached 0.01 significance level (p<0.01, t=7.246 between liver and spleen, 7.769 between liver and small intestine, 7.492 between kidney and spleen, 7.740 between kidney and small intestine, and 6.647 between spleen and small intestine respectively).
As mentioned above, the number of porcine EST hit(s) per human gene was used as an indicator for gene expression activity analysis by setting a certain criterion. By the criterion 3 EST hits per gene (approximately 2,819x0.001), it was found there were 60 BAGs expressed in the major internal organs of the pig, including 6 in the liver, 0 in the kidney, 36 in the spleen, and 18 in the small intestine respectively (Table 4).
As mentioned above, the number of porcine EST hit(s) per human gene was used as an indicator for gene expression activity analysis by setting a certain criterion. By the criterion 3 EST hits per gene (approximately 2,819x0.001), it was found there were 60 BAGs expressed in the major internal organs of the pig, including 6 in the liver, 0 in the kidney, 36 in the spleen, and 18 in the small intestine respectively (Table 4).
Here, about 34,000 human gene cDNA sequences were collected. Of the sequences identified, 33,308 are coding genes distributed in the following Homo sapiens chromosomes (HSC): 3,000 on HSC1, 2,461 on HSC2, 1,872 on HSC3, 1,582 on HSC4, 1,766 on HSC5, 1,847 on HSC6, 1,766 on HSC7, 1,395 on HSC8, 1,368 on HSC9, 1,425 on HSC10, 1,904 on HSC11, 1,557 on HSC12, 777 on HSC13, 1,057 on HSC14, 1,152 on HSC15, 1,258 on HSC16, 1,439 on HSC17, 640 on HSC18, 1,644 on HSC19, 828 on HSC20, 386 on HSC21, 675 on HSC22, 1,315 on HSCX and 195 on HSCY. As for the ESTs sequence resources in the major internal organs of the pig, currently there are a certain amount of cDNA sequences derived from individual or pooled cDNA libraries of liver, kidney, spleen, small intestine and so on.
Of the sequences identified, 33,308 are coding genes distributed in the following Homo sapiens chromosomes (HSC): 3,000 on HSC1, 2,461 on HSC2, 1,872 on HSC3, 1,582 on HSC4, 1,766 on HSC5, 1,847 on HSC6, 1,766 on HSC7, 1,395 on HSC8, 1,368 on HSC9, 1,425 on HSC10, 1,904 on HSC11, 1,557 on HSC12, 777 on HSC13, 1,057 on HSC14, 1,152 on HSC15, 1,258 on HSC16, 1,439 on HSC17, 640 on HSC18, 1,644 on HSC19, 828 on HSC20, 386 on HSC21, 675 on HSC22, 1,315 on HSCX and 195 on HSCY.
3% of the total number of human genes) which had high similarities to the ESTs sequences expressed in the major internal organs of pig. Of these 2,100 genes, 128 genes were expressed in liver (about 6.10% of the total 2,100 genes), 81 in kidney (3.86%), 780 in spleen (37.14%) and 1,423 in small intestine (67.76%) respectively. There were genes co-expressed in these tissues (Table 2).
3% of the total number of human genes) which had high similarities to the ESTs sequences expressed in the major internal organs of pig. Of these 2,100 genes, 128 genes were expressed in liver (about 6.10% of the total 2,100 genes), 81 in kidney (3.86%), 780 in spleen (37.14%) and 1,423 in small intestine (67.76%) respectively. There were genes co-expressed in these tissues (Table 2).
, 1999). The results here showed that an in silica study could be credible and efficient for detecting gene expression profiles in a certain tissue, and it also further confirmed that there were many common aspects in gene expression between humans and pigs. Thus there was relative high conservation in phylogenesis between them.
The expression information of BAGs showed that the expression activities of human genes were apparently different in the major internal organs of the pig. The results indicated it was a efficient way to discover genes preferentially expressed in some tissues by detecting the basic active genes (BAGs), and it was also a brand-new way to discover new genes. On the other hand, we could rapidly acquire the full length cDNA sequence of a certain gene by overlapping fragments assembling of the corresponding ESTs sequences.
(2001) used this protein for microchannel bioreactor study and this doubtlessly gave us some inspiration in exploiting these genes expressed in the internal organs. The top-ranking BAGs expressed in the spleen included CAP1, HLA-DRB3, WBP4, C1QG LOC152059, LOC219398, HLA-DPB1, and LOC221518 (the latter 5 are function-unknown genes). CAP1 encodes adenylate cyclase-associated protein 1.
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