Osteoblast differentiation of rabbit adipose-derived stem cells by polyethylenimine-mediated BMP-2 gene transfection in vitro.

The aim of this study was to examine the efficiency of polyethylenimine-mediated transfection of the human bone morphogenetic protein-2 (BMP-2) gene into rabbit adipose-derived stem cells (ADSCs), and its effect on osteoblast differentiation. Adipose tissue was isolated from the necks of adult Japanese white rabbits and cultured in vitro to obtain ADSCs. Gene delivery of BMP-2 was mediated by polyethylenimine and stable transformants were selected by G-418. The expression of BMP-2 mRNA was confirmed by reverse transcription-polymerase chain reaction, and of the BMP-2 protein by ELISA. Osteocalcin and collagen type I were detected by western blot and by an alkaline phosphatase kit. Alizarin red S stain was also utilized to examine osteogenesis. The non-transfected group was considered as a control. In this study, we successfully derived ADSCs from rabbit adipose tissue. Through passages 3-6, the expression of CD29 and CD44 gradually increased, whereas the expression of CD34 and CD45 gradually decreased. Both mRNA and protein expression of BMP-2 were confirmed following polyethylenimine-mediated BMP-2 gene delivery. In addition, the expression of alkaline phosphatase, osteocalcin, and collagen type I was found to be upregulated and alizarin red S staining was positive in transfected ADSCs, indicating BMP-2-induced osteogenesis. Therefore, this study determined that polyethylenimine was able to mediate BMP-2 gene delivery and induce osteogenic differentiation of ADSCs.

Molecular cloning and expression profile of an ATP-binding cassette (ABC) transporter gene from the hemipteran insect Nilaparvata lugens.

The ATP-binding cassette (ABC) transporters belong to a large superfamily of proteins that have important physiological functions in all living organisms. In insects, ABC transporters have important functions in the transport of molecules, and are also involved in insecticide resistance, metabolism, and development. In this study, the Nilaparvata lugens Stal (Hemiptera: Delphacidae) ABCG (NlABCG) gene was identified and characterized. The complete mRNA sequence of NlABCG was 2608-bp long, with an open reading frame of 2064 bp encoding a protein comprised of 687 amino acids. The conserved regions include three N-glycosylation and 34 phosphorylation sites, as well as seven transmembrane domains. The amino acid identity with the closely related species Acyrthosiphon pisum was 42.8%. Developmental expression analysis using quantitative real-time reverse transcriptase PCR suggested that the NlABCG transcript was expressed at all developmental stages of N. lugens. The lowest expression of NlABCG was in the 1st instar, and levels increased with larval growth. The transcript profiles of NlABCG were analyzed in various tissues from a 5th instar nymph, and the highest expression was observed in the midgut. These results suggest that the sequence, characteristics, and expression of NlABCG are highly conserved, and basic information is provided for its functional analysis.

Isolation, characterization, and cross-transferability of microsatellite markers from the whitebacked planthopper (Sogatella furcifera).

The whitebacked planthopper Sogatella furcifera (Horváth) (Hemiptera: Delphacidae) is one of the most harmful pests of rice. In this study, 18 polymorphic microsatellite markers were developed from S. furcifera genomic libraries using the fast isolation by amplified fragment length polymorphism of sequences containing repeat protocols. Microsatellite polymorphism was investigated using 32 individuals from one natural population. These 18 simple sequence repeat markers showed a number of alleles that ranged from 3 to 15 and had observed and expected heterozygosities that ranged from 0.094 to 0.871 and from 0.148 to 0.924, respectively. The high cross-species transferability of these markers was evaluated in three other planthopper species: Nilaparvata lugens, N. muiri China, and N. bakeri (Muir). These microsatellite markers will provide powerful tools for population genetic and ecological studies of this pest and its related species in the future.

Two whitebacked planthopper resistance genes in rice share the same loci with those for brown planthopper resistance.

The whitebacked planthopper (WBPH), Sogatella furcifera, and brown planthopper (BPH) Nilaparvata lugens Stål are important sucking insects of rice (Oryza sativa L.) crops throughout the world. Rice 'B5', which has derived its resistance genes from the wild rice O. officinalis Wall ex Watt, is a line that is highly resistant to both WBPH and BPH. Previously, two resistance genes against BPH, Qbp1, and Qbp2 in 'B5' had been mapped onto chromosome 3 and chromosome 4, respectively. In this study, we employed a mapping population composed of 187 recombinant inbred lines (RILs), produced from a cross between 'B5' and susceptible variety 'Minghui63', to locate the WBPH and BPH resistance genes. A RFLP survey of the bulked extremes from the RIL population identified two genomic regions, one on chromosome 3 and the other on chromosome 4, likely containing the resistance genes to planthoppers. QTL analysis of the RILs further confirmed that two WBPH resistance genes were mapped on the same loci as Qbp1 and Qbp2, using a linkage map with 242 molecular markers distributed on 12 rice chromosomes. Of the two WBPH resistance genes, one designated Wbph7(t) was located within a 1.1-cM region between R1925 and G1318 on chromosome 3, the other designated Wbph8(t) was within a 0.3-cM region flanked by R288 and S11182 on chromosome 4. A two-way analysis of variance showed that two loci acted independently with each other in determining WBPH resistance. The results have significant implications in studying the interactions between sucking insects and plants and in breeding programs of resistance to rice planthoppers.