PURPOSE: The list of genetically engineered crops is growing. Traits introduced into these crops include insect protection, delayed ripening, virus resistance, modified nutritional composition, herbicide tolerance etc. Most traits introduced into crops result from the expression of new proteins. FAO/WHO organised joint expert consultations had recommended that substantial equivalence be an important component in the safety assessment of GMO plants for human consumption. As the first step to assess the allergenic potential of GMO food, the immunological and physicochemical characterization is needed. METHODS: We made crude extract from GMO soybean, wild soybean, curd and soy milk and performed SDS-PAGE. After acidification with HCl, the samples were divided to globulin and whey. To evaluate the changes of protein composition, the samples were heated or added with pepsin. PCR with primer coding 35S-promotor, NOS-terminator, and EPSPS gene were performed respectively for detection of GMO component. RESULTS: Although there was difference in protein composition in SDS-PAGE of GMO and wild soybean, the same protein bands are observed in globulin fraction after acidification. The heating made difficult to see the protein distribution exactly. After adding of pepsin the same bands-20 kD, 37 kD, and 68 kD-were preserved in GMO and wild soybeans. The 3 PCR procedures showed same results that GMO soybean and some curd included GMO component. CONCLUSION: There were no definite differences between GMO and wild soybeans in respect to immunologic and physicochemical characteristics. To assess the allergenicity of GMO food, the more researches including in vitro and in vivo immunoassay are needed.
Clinical Coding ; Electrophoresis, Polyacrylamide Gel ; Excitatory Postsynaptic Potentials ; Food, Genetically Modified ; Heating ; Hot Temperature ; Humans ; Immunoassay ; Insects ; Joints ; Organisms, Genetically Modified ; Pepsin A ; Plants, Genetically Modified ; Polymerase Chain Reaction ; Referral and Consultation ; Soy Milk ; Soybeans*

This study was conducted to evaluate the safety of herbicide-resistant rice, a genetically modified organism (GMO) developed by the Rural Development Administration, in Sprague-Dawley rats. The nutrient content of herbicide-resistant polished and brown cooked rice was compared with that of conventional Ilpum polished and brown cooked rice to assess composition equivalence. Compositional analysis was performed to measure proximates, fiber, and minerals before animal feeding. Growing male rats were fed one of the following four diets for six weeks: Ilpum polished cooked rice (IP) and Ilpum brown cooked rice (IB) as a non-GMO and herbicide-resistant polished cooked rice (GP) and brown cooked rice (GB) as a GMO. We checked clinical symptoms (anorexia, salivation, diarrhea, polyuria, anuria, fecal change) every day, food intake, change of body weight twice a week, and serum biochemistry and organ weights after 6 weeks of experimental feeding among the four groups. Nutrient content of the herbicide-resistant rice was similar to that of the non-transgenic control and was within the published range observed for non-transgenic rice. We could not find any significant difference in the above-mentioned items as the index to be checked in the animals fed the GMO. These results suggest that the nutrient content of genetically modified herbicide-resistant rice is compositionally equivalent to that of conventional Ilpum rice and that growing male rats fed herbicide-resistant rice are no different from those fed Ilpum rice, non-GMO for 6 weeks.
Animal Feed ; Animals ; Anuria ; Biochemistry ; Body Weight ; Diarrhea ; Diet ; Eating ; Humans ; Male* ; Minerals ; Organ Size ; Organisms, Genetically Modified ; Polyuria ; Rats* ; Rats, Sprague-Dawley ; Salivation ; Social Planning

Animals ; Bone Marrow Cells ; cytology ; metabolism ; Cell Proliferation ; Interleukin-10 ; genetics ; metabolism ; Mesenchymal Stromal Cells ; cytology ; metabolism ; Organisms, Genetically Modified ; Rats ; Rats, Sprague-Dawley ; Transfection

OBJECTIVETo study the expression of brain-derived neurotrophic factor (BDNF) gene modified bone marrow mesenchymal stem cells (MSC) in the cochlea of drug-deafened guinea pigs and its protection to spiral ganglion cells (SGC).

METHODSGuinea pigs deafened by subcutaneous injection of amikacin were randomly divided into two groups, BDNF gene modified bone marrow MSC were injected into the cochlea through fenestration of scala tympani in the experimental group, while artificial perilymphatic fluid were injected in the control group. Experimental animals were executed at 7 and 28 days post-operation. Expression of BDNF mRNA was examined by quantitate real time RT-PCR, histological images of cochlear sections were analyzed to calculate the cellular density of the SGC, and terminal deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL) was used to identify the apoptotic neurons.

RESULTSThe BDNF expressive level in experimental group was higher than in the control group at 7 d and 28 d post-operation, whose differences were both statistically significant (P < 0.01). And, It showed a higher abundance of ganglion cell numbers, as well as a decreased apoptotic index in experimental group compared with the control group at 7 d and 28 d post-operation, whose differences were all statistically significant (P < 0.01).

CONCLUSIONBDNF gene modified MSC could maintain expression for at least 28 days after transplantation into cochlea of drug deafened guinea pigs, and protect SGC.

Animals ; Bone Marrow Cells ; metabolism ; Brain-Derived Neurotrophic Factor ; genetics ; pharmacology ; Deafness ; chemically induced ; therapy ; Guinea Pigs ; Mesenchymal Stromal Cells ; metabolism ; Organisms, Genetically Modified ; Spiral Ganglion ; drug effects

Various Leishmania species were engineered with green fluorescent protein (GFP) using episomal vectors that encoded an antibiotic resistance gene, such as aminoglycoside geneticin sulphate (G418). Most reports of GFP-Leishmania have used the flagellated extracellular promastigote, the stage of parasite detected in the midgut of the sandfly vector; fewer studies have been performed with amastigotes, the stage of parasite detected in mammals. In this study, comparisons were made regarding the efficiency for in vitro G418 selection of GFP-Leishmania amazonensis promastigotes and amastigotes and the use of in vivo G418 selection. The GFP-promastigotes retained episomal plasmid for a prolonged period and G418 treatment was necessary and efficient for in vitro selection. In contrast, GFP-amastigotes showed low retention of the episomal plasmid in the absence of G418 selection and low sensitivity to antibiotics in vitro. The use of protocols for G418 selection using infected BALB/c mice also indicated low sensitivity to antibiotics against amastigotes in cutaneous lesions.
Amebicides/*pharmacology ; Animals ; Flow Cytometry ; Gentamicins/*pharmacology ; Green Fluorescent Proteins/*chemistry ; Host-Parasite Interactions ; Leishmania mexicana/drug effects/genetics/*growth & development ; Leishmaniasis, Cutaneous/*parasitology ; Luminescent Agents/*chemistry ; Macrophages, Peritoneal/parasitology ; Mice ; Mice, Inbred BALB C ; Organisms, Genetically Modified ; Spectrometry, Fluorescence

This study was aimed to investigate the protective effect of Wit3a gene modification on mouse bone marrow mesenchymal stem cells against the injury induced by Ara-C. The gene-modified MSC steadily expressing Wnt3a were established by adenovirus system. The acute direct damage effects of different concentrations of Ara-C on the unmodified MSC and the gene-modified MSC were assessed by using an in vitro culture system, and the corresponding controls were set. The proliferation and apoptosis of MSC exposed to Ara-C were detected by cell count kit-8 (CCK-8) and flow cytometry. The expression of BCL-2 protein related with cell apoptosis was assayed by Western blot. The results indicated that as compared with unmodified MSC, Ara-C exhibited a less inhibitory effect on the proliferation of gene-modified MSC. There was obvious difference between unmodified MSC and gene-modified MSC (p < 0.05). The proliferation of gene-modified MSC began to recover at 72 hours after removal of Ara-C. However, unmodified MSC showed sustained suppression of proliferation after withdrawal of Ara-C. In apoptosis, the apoptosis rate of gene-modified MSC induced by Ara-C was significantly lower than those of unmodified MSC (p < 0.05). In addition, the expression levels of BCL-2 protein in gene-modified MSC were up-regulated compared with unmodified MSC (p < 0.05). It is concluded that Wnt3a gene modification can significantly mitigate the damage of mouse bone marrow MSC induced by Ara-C.
Animals ; Bone Marrow Cells ; drug effects ; metabolism ; Cytarabine ; adverse effects ; Mesenchymal Stromal Cells ; drug effects ; metabolism ; Mice ; Organisms, Genetically Modified ; Proto-Oncogene Proteins ; metabolism ; Proto-Oncogene Proteins c-bcl-2 ; Wnt3A Protein ; genetics

OBJECTIVEConstruction of human full-length survivin gene-modified DCs vaccine and observe the biological characteristics and the role of laryngeal cancer treatment.

METHODSConstruction of homologous recombination through the full-length survivin-human adenovirus (pAd-sur), and transfection of immature DCs, induced by access to sophisticated cultivate DCs, the proliferation of activated T lymphocytes; DCs vaccine different groups observed in vitro, in vivo biological activity.

RESULTSThe success in bringing human to build a full-length survivin gene recombinant adenovirus vector pAd-sur; get with the typical characteristics of DCs, the expression level of the surface molecules of up to CD1a 46.2%, CD80 64.2%, CD83 80.5%, HLA-DR 84.3%, vaccine for in vitro cell killing rate 51.5%, compared with the control group, the difference was significant (t = 7.1358, P < 0.01). Gene-modified tumor cell vaccine group was 54.9% of apoptosis and necrosis rate of 21.9%, compared with the control group, the difference was significant (t value was 33.0209 and 17.5057, all P < 0.01).

CONCLUSIONHuman full-length survivin gene modified, DCs The expression of surface molecules than the rate of gene-modified DCs without a significant increase, with more to stimulate T lymphocyte proliferation, and promote secretion of gamma-IFN; in vivo, in vitro to promote the ability of anti-laryngeal cancer cells.

Animals ; Cancer Vaccines ; immunology ; Cell Line, Tumor ; Dendritic Cells ; immunology ; Female ; Genetic Vectors ; Humans ; Inhibitor of Apoptosis Proteins ; Laryngeal Neoplasms ; immunology ; prevention & control ; Mice ; Mice, Nude ; Microtubule-Associated Proteins ; genetics ; immunology ; Organisms, Genetically Modified

To construct the recombinant vector Pact-EGFP, the Act-1 core promoter region was amplified from the pUCm-T/Act-1 and subcloned into pEGFP-4.1 vector (derived from pEGFP-N1 with the removal of human cytomegalovirus immediate early promoter), by restriction enzymes Bgl II and Hind III. Transfection of Pact-EGFP vector into Vero cell by liposome indicated that Act-1 core promoter regulated the expression of EGFP gene in lower level in Vero cells. After Pact-EGFP microinjection into the gonad of Caenorhabditis elegans with pRF4 as a gene marker, green fluorescence was detected in the cortex, vice cortex and the pharyngeal of C. elegans. According to the locations, two different transgene lines were separated. The expression level of EGFP expressed in C. elegans was more than that in Vero cell. Some unique motifs might exist in Act-1 core promoter region of C. elegans, which was closely related to the expression level of EGFP. These results lay the foundation for the further research on gene function of parasitic nematodes using C. elegans.
Animals ; Caenorhabditis elegans ; genetics ; metabolism ; Cercopithecus aethiops ; Connexin 43 ; genetics ; Green Fluorescent Proteins ; genetics ; metabolism ; Organisms, Genetically Modified ; Peptide Fragments ; genetics ; Promoter Regions, Genetic ; genetics ; Transcription, Genetic ; Vero Cells

This study was aimed to investigate the homing capacity of CXCR4 overexpressed mesenchymal stem cells (MSC) and their effect on hematopoietic recovery. The 293FT packaging cell line was transfected with the recombinant lentiviral vector LV-CXCR4-IRES-EGFP and LV-IRES-EGFP to produce lentivirus. Mouse MSC were then infected with viral supernatant. Male BALB/c mice were sublethally irradiated and then were injected intravenously with 5×10(5) MSC. General status and survival rate of mice were observed every day. On day 3, 7, 14, 21 and 28, peripheral blood samples were collected to calculate the number of white blood cells (WBC) and red blood cells (RBC), the ratio of reticulocyte to platelet, the number of platelet was detected by flow cytometry. The recovery of bone marrow and spleen was pathologically monitored. The proportion of MSC implantation was analysed by PCR. The results showed that the peripheral blood cells displayed the tendency of firstly increasing and then decreasing to their normal level. Generally, recovery of WBC level was earlier in mice infused with MSC (P < 0.05) . The histopathological examination of spleen and bone marrow showed a faster hematopoietic recovery in CXCR4-MSC group than the other two groups. And the donor MSC could be detected in the recipients on day 7, 14, 21 and 28. It is concluded that infusion of CXCR4-MSC enhances the implantation of hematopoietic stem cells and promotes hematopoietic recovery of the sublethally irradiated mice.
Animals ; Bone Marrow Cells ; cytology ; Genetic Vectors ; Hematopoiesis ; Male ; Mesenchymal Stem Cell Transplantation ; Mesenchymal Stromal Cells ; cytology ; Mice ; Mice, Inbred BALB C ; Organisms, Genetically Modified ; Radiation Injuries, Experimental ; therapy ; Receptors, CXCR4 ; genetics ; Transfection ; Whole-Body Irradiation

Iro system and temperature-sensitive hemagglutinin (Tsh) genes were identified by suppression subtractive hybridization (SSH) and selective capture of transcribed sequences (SCOTS). To get more insights in the distribution and the occurrence of the iroC and tsh genes, we examined 243 avian E. coli strains for the presences of the these genes. Among 243 avian E. coli isolates, iroC gene was present in 84.4% strains (205/243). Of the 205 iroC-positive isolates, iroC gene was found in 184 (89.8%), 18(8.8%) and 3 (1.5%) isolates with high, intermediate and low pathogenicity, respectively. Of the 167 tsh-positive isolates, tsh gene was detected in 146 (87.4%), 21 (12.6%) and 0 (0%) isolates with high, intermediate and low pathogenicity, respectively. Among tsh-positive isolates, 89.5 to 100% of the highly pathogenic isolates of O1, O2 or O78 serogroups had the tsh gene, while 53.3% of the highly pathogenic isolates of non-O1, O2 and O78 serogroups had the tsh gene (P<0.01). Suicide vectors for deletion of the iroBCDEN or tsh genes were constructed as follows. The 715-bp fragments of iroB and 603-bp fragment of the iroN were generated by PCR respectively. Both of these two fragments together with EGFP gene were cloned into pUC18, termed pUC18-iroBNEGFP. A resultant suicide vector containing the iroB-EGFP-iroN fragment was obtained and named pMEG375-iroBNEGFP. Similarly, both of the 685-bp fragment of tshF and the 692-bp fragment of the tshR together with gentamycin gene were cloned into pUC18, resulting in pUC18-tshFRGm. A resultant suicide vector containing the tshF-Gm-tshR fragment was named pMEG375-tshFRGm. Mutant derivatives of strain E037 were generated by allelic replacements and were named E037(Deltairo), E037(Deltatsh) and E037(DeltairoDeltatsh). The 50% lethal dose (LD50) of E037, E037(Deltairo), E037(Deltatsh) and E037(DeltairoDeltatsh) in commercial day-old chickens experimentally inoculated via intratrachea were determined to be 10(5.6), 10(8.4), 10(9.0) and 10(9.5)CFU, respectively. In the chicken challenging model, the mutants were tested to determine the individual role of this system for virulence and persistence in chickens. The result suggested that Iro system and Tsh were important in the pathogenicity of APEC.
Adhesins, Escherichia coli ; genetics ; Animals ; Chickens ; Escherichia coli ; genetics ; pathogenicity ; Escherichia coli Infections ; microbiology ; veterinary ; Genes, Bacterial ; genetics ; Mutation ; Nucleic Acid Hybridization ; methods ; Organisms, Genetically Modified ; Poultry Diseases ; microbiology ; Transformation, Genetic ; Virulence Factors ; genetics