Plant Expression Vector Containing Antifreeze Peptide Gene and Its Applications in Transgenic Plants, Secretory Expression in Trichoderma Viride and a Study on Intron Function of AFP Gene
Molecular Cloning of 5' Flanking Region and Introns of 20 a Hydroxysteroid Dehydrogenase Gene,Structure Analysis and Measurement on their Activities of Regulation Gene Expression
The full length of Lac1 gene was 2338bp, with 10 introns and 11 exons. Introns ranged in size from 51 to 76bp, and its sequence complied with the rule of “5'-gt” and “ag-3'”.
To generate transgenic porcine which expresses human serum albumin(HSA), the HSA gene targeting vector was constructed with HSA cDNA as the gene of interestand partial porcine serum albumin (PSA) gene as homologous arms which respectively were 7.2kb 5′ regulation sequence and 2.8 kb genomic sequence from the first intron to the fourth intron.
The full length of chalcone synthase gene of Glycine max L(GMCHS)was cloned from the genomic DNA,and the two exons(GMCHSexon)with length of 1170bp in total were jointed by SOE-PCR method. The DNA sequence analysis showed the 390 amino acid coding area has 97% sequence similarity to the published GmCHS gene(AY237728).
Through sequencing and analyzing by method of bioinformatics: SDIFN-αORF includes 576 nucleotides, encoding 191 amino acids. Molecular weight is about 21.64kDa, front 28 amino acide is signal peptide, and the cleavage site lies in 28-29 amino acide(ANA-FS).
A pair of primers, designed from conserved regions of plant soluble acid invertase gene, was used to amplify 1096bp fragment by polymerase chain reaction (PCR). The fragment was cloned into pUCm-T, and then sequenced.
Furthermore, characteristics of EID, such as intron phase, distribution of different splice sites, and the relationship between genome size and intron proportion or intron density, have been studied.
The exon-intron structure of these genes has been analyzed in zebrafish Danio rerio, loach Misgurnus fossilis, fugu Takifugu rubripes, and Nile puffer Tetraodon fahaka.
This is supported by high sequence similarity between mlc1 and mlc3 as well as by the exon-intron structure of these genes and their localization on different chromosomes.
The result indicated that the sequence was 1,813 bp (BamHI and SacI were introduced at the 5' and 3' end) including 7 extrons and 6 introns, coding 238 amino acids.
Studies on the statistical characteristics of EID show that there were 103, 848 genes, 478,484 introns, and 582,332 exons, with an average of 4.61 introns and 5.61 exons per gene.
Results of the statistical analysis on the data from nine model species showed that in eukaryotes, higher species do not necessarily have more introns or exons in a gene than lower species.
As shown by nitrocellulose filtration assays with RNA fragments transcribed from various regions of the human ribosomal protein (rp) S26 gene, proteins of the 40S ribosome subunit bind to the first intron of the rpS26 pre-mRNA.
Small-subunit proteins did not affect the efficiency of in vitro splicing of a pre-mRNA fragment corresponding to the first intron, second exon, second intron, and a part of the third exon of the rpS26 gene.
Immunoassays with specific antibodies showed that rpS26 contained in the nuclear extract of HeLa cells binds to the first intron of its pre-mRNA and, less efficiently, to its mRNA.