Biomedical subjects

Qin Chen

Publications and source records attributed to Qin Chen.

At least 37 records · Page 2Linked to original sources

PubMed2005

Transfection and expression of exogenous gene in laying hens oviduct in vitro and in vivo.

To examine whether or not the regulatory sequence of chicken ovalbumin gene can drive transgene expression specifically in hen oviduct, the authors constructed an oviduct-specific expression vector (pOV), containing 3.0 kilobases (kb) of the 5'-flanking sequence and 3.0 kb of the 3'-flanking sequence of the chicken ovalbumin gene. Jellyfish green fluorescence protein (EGFP) reporter gene and bacterial LacZ reporter gene were respectively inserted into the downstream of the 5'-regulatory region. The recombinants were named as pOVEGFP and pOVLacZ. Two transfer systems, in vitro and in vivo, were used to verify the function of the vector. In vitro, the plasmid DNA pOVEGFP and pEGFP-N1 were transfected respectively by the polyethyleneimine procedure into the primary chicken oviduct epithelium (PCOE) and fibroblasts cells isolated from laying hens. In vivo, the recombinant vector pOVLacZ was injected into egg-laying hens via wing vein and the tissues were collected for RT-PCR analysis. The results showed that expression of pEGFP-N1 was achieved at low level in oviduct epithelial cells and at high level in fibroblasts, but that the recombinant vector was not expressed in both cells. RT-PCR analysis showed that the LacZ gene was transcribed in the oviduct, but not in the heart, liver, kidney and spleen of the injected hens. Accordingly, the beta-galactosidase activity was only detected in the oviduct magnum (116.7 mU/ml) and eggs (16.47 mU/ml). These results indicated that the cloned regulation regions of chicken ovalbumin gene could drive exogenous gene expression specifically in the oviducts of hens. In vivo gene injection via wing vein may serve as a rapid production system of recombinant proteins in chicken eggs. In addition, the cultured primary oviduct cells from laying hens were not efficient temporary expression systems for analyzing the function of regulating elements of ovalbumin gene.

Animals↗

PubMed2005

Gene therapy for Parkinson's disease: progress and challenges.

Therapy for Parkinson's disease (PD), a common neurological disorder characterized by pathological degeneration of the nigrostriatal dopaminergic system, remains unsatisfactory. Gene therapy is considered one of the most promising approaches to developing a novel effective treatment for PD. Among the numerous candidate genes that have been tested as therapeutic agents, those encoding tyrosine hydroxylase, guanosine triphosphate cyclohydrolase I and aromatic L-amino acid decarboxylase all boost dopamine production, while glial cell line-derived neurotrophic factor promotes the survival of dopaminergic neurons and is generally believed to possess the greatest potential for successful restoration of the dopaminergic system. The genes encoding vesicular monoamine transporter-2 and glutamic acid decarboxylase have also produced therapeutic effects in animal models of PD. Both viral and non-viral vectors, each with its particular advantages and disadvantages, have been used to deliver these genes into the brain. Whether or not regulatable expression systems are essential to successful gene therapy for PD remains a critical issue in the clinical application of this emerging treatment. Here we review the current status of gene therapy for PD, including the application of control systems for transgene expression in the brain.

Animals↗

PubMed2005

[Clinical study of wind-warm and pulmonary heat syndrome treated with integrated traditional Chinese and Western medicine].

OBJECTIVE: To investigate the clinical effectiveness of Toubiao Qingfei (expelling exterior evil and clearing lung) Decoction (TBQFD) on wind-warm and pulmonary heat syndrome. METHODS: Forty-six subjects were randomized into treatment group and control group. Patients in the control group were treated with Western medicines, while patients in the treatment group were treated with Western medicines and TBQFD. The therapeutic effects and improvement of symptoms in both groups were observed. RESULTS: The durations of fever, cough and absorption of pulmonary inflammatory focus in the treatment group were 1.52, 3.52 and 6.25 days respectively, which were significantly shorter than those in the control group (P<0.05). CONCLUSION: TBQFD can improve the symptoms, such as fever and cough, and can promote the absorption of infection and shorten the clinical course of wind-warm and pulmonary heat syndrome.

Adult↗

PubMed2005

[Experimental study on the mechanism of the apoptosis of leukemic cells induced by valproic acid].

OBJECTIVE: To overcome the drug-resistance of tumor cells is one of the methods of improving the therapeutic results. Histone deacetylase inhibitors (HDACIs) is a novel class of chemotherapeutic agents which can induce apoptosis of tumor cells. Valproic acid (VPA) is a common drug used in the treatment of epilepsy. It has been shown that VPA has a marked HDACIs effect at the pharmaceutical level, and can induce the differentiation and apoptosis of transformed cells. But the mechanism of its effect has not been clarified. The aim of this study was to investigate the mechanism of VPA in inducing the apoptosis of leukemic cells at molecular level. METHODS: The cell lines U937, Jurkat clone E6 - 1 (Jurkat) and BALL-1 were cultured in RPMI 1640 medium containing 20% calf serum, then divided into three groups (control group, 1 mmol/L VPA group and 1 mmol/L VPA + 1 micromol/L Pan-caspase inhibitor zVAD-fmk group). At 72 hours after the treatment, the cells were double stained with Aunexin and PI (propidium iodide) and then were analyzed with the flow cytometry (FCM) to detect apoptosis. Before and after treatment with VPA the mean fluorescence index (MFI) of Bcl-2, Bax, Bcl-xl and the levels of caspase 8, 9 and 3 were also detected with the FCM. The changes of P(44/42) mitogen activating protein kinase (MAPK) and phosphorylated P(44/42) MAPK were determined by Western blotting. RESULTS: Seventy-two hours after the treatment, 1 mmol/L VPA induced apoptosis of U937 and Jurkat. The apoptotic rate of U937 was (75.78 +/- 4.20)% and that of Jurkat was (53.50 +/- 5.87)% (P < 0.01, vs. control group); zVAD-fmk could fully inhibit the apoptosis of U937, and the apoptotic rate was (2.89 +/- 0.36)%; while it could partly inhibit the apoptosis of Jurkat, and the apoptotic rate was (15.38 +/- 1.40)% (P < 0.01). 1 mmol/L VPA could not induce the apoptosis of BALL-1 which had a high expression level of Bcl-2. The MFI of Bcl-2, Bax and Bcl-xl in these three cell lines did not change significantly with VPA (P > 0.05). After treatment with VPA, the level of caspase 3 in U937 increased from (14.09 +/- 1.19)% to (32.30 +/- 2.47)%, and caspase 8 from (4.58 +/- 1.41)% to (86.47 +/- 3.26)% (P < 0.01), but there was no significant change in caspase 9 [(13.25 +/- 3.11)% and (10.95 +/- 1.30)%]. In Jurkat, the level of caspase 3 increased from (12.01 +/- 1.63)% to (35.56 +/- 0.27)%, and caspase 9 from (13.89 +/- 1.71)% to (75.89 +/- 4.08)% (P < 0.01 for both); no significant change was observed for caspase 8 [(5.94 +/- 1.38)% and (5.44 +/- 0.72)%]. In BALL-1, there was a slight decline in caspase 3 (P < 0.05). With the effect of VPA, levels of P(44/42) MAPK and phosphorylated P(44/42) MAPK decreased in all three cell lines (P < 0.01). CONCLUSION: VPA could induce apoptosis of U937 through the activation of caspase 3 and 8; and it induced the apoptosis of Jurkat involving the activation of caspase 3 and 9. P(44/42) MAPK pathway also plays an important role in this course. VPA induced apoptosis of these cell lines without the alteration of Bcl-2, Bax and Bcl-xl. High level of Bcl-2 could antagonize the effect of VPA.

Apoptosis↗

PubMed2004

Inhibition of lens fiber cell morphogenesis by expression of a mutant SV40 large T antigen that binds CREB-binding protein/p300 but not pRb.

Simian virus (SV) 40 large T antigen can both induce tumors and inhibit cellular differentiation. It is not clear whether these cellular changes are synonymous, sequential, or distinct responses to the protein. T antigen is known to bind to p53, to the retinoblastoma (Rb) family of tumor suppressor proteins, and to other cellular proteins such as p300 family members. To test whether SV40 large T antigen inhibits cellular differentiation in vivo in the absence of cell cycle induction, we generated transgenic mice that express in the lens a mutant version of the early region of SV40. This mutant, which we term E107KDelta, has a deletion that eliminates synthesis of small t antigen and a point mutation (E107K) that results in loss of the ability to bind to Rb family members. At embryonic day 15.5 (E15.5), the transgenic lenses show dramatic defects in lens fiber cell differentiation. The fiber cells become post-mitotic, but do not elongate properly. The cells show a dramatic reduction in expression of their beta- and gamma-crystallins. Because CBP and p300 are co-activators for crystallin gene expression, we assayed for interactions between E107KDelta and CBP/p300. Our studies demonstrate that cellular differentiation can be inhibited by SV40 large T antigen in the absence of pRb inactivation, and that interaction of large T antigen with CBP/p300 may be enhanced by a mutation that eliminates the binding to pRb.

Animals↗

PubMed2004

Subtypes of BK virus prevalent in Japan and variation in their transcriptional control region.

BK polyomavirus (BKV) is ubiquitous in the human population, infecting children without obvious symptoms, and persisting in the kidney in a latent state. In immunosuppressed patients, BKV is reactivated and excreted in urine. BKV isolates have been classified into four subtypes (I-IV) using either serological or genotyping methods. To elucidate the subtypes of BKV prevalent in Japan, the 287 bp typing region in the viral genome was PCR-amplified from urine samples of 45 renal transplant (RT) and 31 bone-marrow transplant (BMT) recipients. The amplified fragments were subjected to a phylogenetic or RFLP analysis to determine the subtypes of BKV isolates in urine samples. Subtypes I, II, III and IV were detected, respectively, in 70-80, 0, 2-3 and 10-20 % of the BKV-positive patients in both patient groups. This pattern of distribution was virtually identical to patterns previously demonstrated in England, Tanzania and the United States, suggesting that BKV subtypes are distributed similarly in various human populations. Furthermore, transcriptional control regions (TCRs) were PCR-amplified from the urine samples of 25 RT and 20 BMT recipients, and their nucleotide sequences were determined. The basic TCR structure (the so-called archetype configuration) was observed in most isolates belonging to subtypes I, III and IV (subtype II isolates were not available), albeit with several nucleotide substitutions and a few single-nucleotide deletions (or insertions). Only three TCRs carried extensive sequence rearrangements. Thus, it was concluded that the archetypal configuration of the BKV TCR has been conserved during the evolution of BKV.

BK Virus↗

PubMed2004

The transfer and characterization of resistance to common root rot from Thinopyrum ponticum to wheat.

Common root rot, caused by Cochliobolus sativus (Ito and Kurib) Drechs. ex Dastur, is a major soil-borne disease of spring and winter wheat (Triticum aestivum L. em Thell.) on the Canadian prairies. Resistance to common root rot from Thinopyrum ponticum (Podp.) Liu and Wang was transferred into wheat via crossing with Agrotana, a resistant wheat - Th. ponticum partial amphiploid line. Evaluation of common root rot reactions showed that selected advanced lines with blue kernel color derived from a wheat x Agrotana cross expressed more resistance than the susceptible T. aestivum 'Chinese Spring' parent and other susceptible wheat check cultivars. Cytological examination revealed 41 to 44 chromosomes in the advanced lines. Genomic in situ hybridization, using total genomic DNA from Pseudoroegneria strigosa (M. Bieb) A. Löve (St genome) as a probe, demonstrated that the blue kernel plants had two pairs of spontaneously translocated J-Js and Js-J chromosomes derived from the J and Js genome of Th. ponticum. The presence of these translocated chromosomes was associated with increased resistance of wheat to common root rot. The lines with blue aleurone color always had a subcentromeric Js-J translocated chromosome. The subtelocentric J-Js translocated chromosome was not responsible for the blue kernel color. The genomic in situ hybridization analysis on meiosis revealed that the two spontaneous translocations were not reciprocal translocations.

Ascomycota↗

PubMed2004

Distinct capacities of individual E2Fs to induce cell cycle re-entry in postmitotic lens fiber cells of transgenic mice.

PURPOSE: Inactivation of the retinoblastoma gene in human retinoblasts or mouse lens fiber cells causes inappropriate cell cycle entry, presumably as a consequence of elevated activity of the E2F transcription factors. Although E2Fs are known to be critical regulators of the cell cycle, it is still unclear whether family members E2F3a, E2F4 or E2F5 are individually capable of inducing cell cycle entry in vivo. In this study, we designed experiments to test whether lens-specific expression of these E2F family members would induce postmitotic fiber cells to re-enter the cell cycle. METHODS: Transgenic mice were generated by microinjection of constructs that contained E2F cDNAs (E2F3a, E2F4 or E2F5) linked to the mouse lens-specific alphaA-crystallin promoter. The mice were characterized by histology, in situ hybridization, immunohistochemistry, BrdU incorporation, TUNEL assay and Western blots. RESULTS: E2F3a expression was sufficient to induce cell cycle entry in lens fiber cells. Cell cycle re-entry was accompanied by apoptotic cell death resulting in microphthalmia. E2F4 expression stimulated a modest level of cell cycle re-entry, but the transgenic lenses remained normal in size and did not show significant apoptosis. Transgenic mice expressing E2F5 did not show lens defects. In both the E2F3a and E2F4 transgenic lenses, cyclin A2 and cyclin B1 expression were upregulated. Phosphorylated histone H3, a marker for mitosis, was detected in the E2F3a fiber cells. Western blots showed that both p53 and p73alpha were upregulated in the E2F3a lenses. However, expression of p21, a well-known p53 target gene, was not activated, suggesting that p73alpha might be responsible for inducing apoptosis and blocking unregulated proliferation in lens cells overexpressing E2F3a. CONCLUSIONS: E2F3a and E2F4, but not E2F5, function to induce cell cycle entry, although E2F4 has more modest activity. E2F3a may induce cell death primarily through activation of p73alpha.

Animals↗

PubMed2004

Chick delta1-crystallin enhancer influences mouse alphaA-crystallin promoter activity in transgenic mice.

PURPOSE: Both the -366/+43 and the -282/+43 mouse alphaA-crystallin (or alphaA) promoters have been effective at driving transgene expression in lens fiber cells, but not in lens epithelium. Because the chick delta1-crystallin gene is expressed in lens epithelial cells, an enhancer was borrowed from this gene and linked to the alphaA promoter. This heterogenic enhancer/promoter construct was tested in transgenic mice to see whether it was active in both lens epithelium and fiber cells while retaining lens specificity. METHODS: The third intron of the chick delta1-crystallin gene, which contains a lens enhancer element, was added to the 5' end of the mouse alphaA promoter. We refer to this chimeric regulatory element as the deltaenalphaA promoter. To test its activity, we inserted coding sequences for five different genes. Transgenic mice were generated by pronuclear microinjection. Transgene expression patterns were analyzed by either X-gal staining, in situ hybridization or immunohistochemical staining. RESULTS: When deltaenalphaA-lacZ transgenic embryos were stained with X-gal at embryonic day (E)11.5, beta-galactosidase activity was detected only in the eye. Histologic sections of the stained embryos revealed that lacZ was expressed exclusively in the lens, in both epithelial and fiber cells. Transgenic mice were also generated using either the original alphaA- or the new deltaenalphaA promoter linked to an insulin cDNA. In situ hybridizations confirmed that the short alphaA promoter targeted prenatal insulin expression specifically to the lens fiber cells, whereas the deltaenalphaA promoter was active in both lens epithelial and fiber cells. Developmental studies of the deltaenalphaA-insulin mice showed that the deltaenalphaA promoter became active at the lens pit stage and remained active in all lens cells, even at postnatal ages. The deltaenalphaA promoter also successfully directed expression of SV40 T-antigen (TAg), human E2F2, and dominant negative Sprouty2 (dn-Spry2) genes to lens epithelial and fiber cells. The lens specificity of the deltaenalphaA promoter was maintained in minigenes with different types of introns and polyadenylation signals. CONCLUSIONS: A new lens-specific regulatory element was generated-the deltaenalphaA promoter, which can drive high levels of transgene expression in both lens epithelium and fiber cells throughout development. This modified promoter can be used for future transgenic studies of signal transduction and cell cycle regulation in lens epithelial cells.

Animals↗

PubMed2004

Tonoplast H+-ATPase activity in barley roots is regulated by ATP and pyrophosphate contents under NaCl stress.

The changes in ATP and PPi contents in roots under NaCl stress, and the effect of PPi on tonoplast H(+)-ATPase activity were studied with two barley cultivars differing in salt tolerance. The results showed that tonoplast H(+)-ATPase activity in salt-tolerant barley cultivar "Tanyin 2" roots increased obviously under NaCl 200 mmol/L (containing half-strength Hoagland solution) for 2 d, and decreased gradually. Tonoplast H(+)-PPase activity always decreased under NaCl stress. For salt-sensitive barley cultivar "Kepin 7", both tonoplast H(+)-ATPase and H(+)-PPase activities decreased during NaCl stress, as compared with control(grown in half-strength Hoagland solution), ATP content of "Tanyin 2" roots increased under NaCl stress for 2 d, and decreased on 4 d after NaCl stress, as compared with control. In "Kepin 7" roots, ATP accumulation was inhibited during NaCl stress. Pyrophosphate (PPi) content increased slightly both in "Tanyin 2" and "Kepin 7" roots under NaCl stress. Tonoplast H(+)-ATPase activity was inhibited competitively by PPi. These results indicated that the increase in rate of ATP accumulation might be essential for the stimulation of tonoplast H(+)-ATPase activity induced by NaCl. The decrease in ATP content and the inhibition of PPi might result in the decrease of tonoplast H(+)-ATPase activity in barley roots under NaCl stress.

Adenosine Triphosphate↗

PubMed2003

Wnt5a inhibits B cell proliferation and functions as a tumor suppressor in hematopoietic tissue.

Wnt5a is a member of the Wnt family of secreted glycoproteins that play essential organizing roles in development. Similar to other Wnt members, Wnt5a can upregulate cell proliferation and has been proposed to have oncogenic function. Here we report that Wnt5a signals through the noncanonical Wnt/Ca++ pathway to suppress cyclin D1 expression and negatively regulate B cell proliferation in a cell-autonomous manner. Wnt5a hemizygous mice develop myeloid leukemias and B cell lymphomas that are clonal in origin and display loss of Wnt5a function in tumor tissues. Furthermore, analysis of human primary leukemias reveals deletion of the WNT5A gene and/or loss of WNT5A expression in a majority of the patient samples. These results demonstrate that Wnt5a suppresses hematopoietic malignancies.

Animals↗

PubMed2003

Atopic manifestations, breast-feeding protection and the adverse effect of DDE.

Despite the numerous studies on the possible protective effect of breast feeding against the onset of atopic manifestations during childhood, this issue remains controversial. As part of an environmental epidemiological study, we investigated whether different blood concentrations of dichlorodiphenyl-dichloroethylene (DDE) modified the protective effect of breast feeding against atopic manifestations in 338 children. DDE concentration, duration of breast feeding and manifestation of atopic disorders were measured in 1994-95 at age 7-8 years. Information gathered on asthma, atopic eczema and hay fever was based on questionnaire data. We measured the total serum concentration of immunoglobulin E (IgE) and specific IgE levels against inhalant allergens. In 1997, we also determined bronchial hyper-reactivity with a hypertonic saline challenge test. To estimate odds ratios from our cross-sectional analysis, we applied logistic regressions, controlling for confounders. Breast feeding had a protective effect on the two asthma variables (e.g.> 12 weeks breast feeding for doctor-diagnosed asthma, OR = 0.32 [95% CI 0.11, 0.87]; for 'ever' asthma, OR = 0.13 [95% CI 0.02, 0.68]), but not on bronchial hyper-reactivity, hay fever, atopic eczema or the two IgE variables. The protective effect became stronger in children with DDE blood levels below the median of 0.29 micro g/L (e.g. doctor-diagnosed asthma,> 12 weeks breast feeding, OR = 0.24 [95% CI 0.06, 0.95]). Also, for specific IgE against inhalant allergens, the association gained statistical significance. For children with a DDE concentration of 0.29 micro g/L and higher, breast feeding did not show a significant protective effect. Our results suggest that contaminants such as DDE may modify the protective effect and may have contributed to inconsistent findings on the protective effect of breast feeding in previous studies. We recommend determining levels of breast milk contaminants in children when assessing the impact of breast feeding on atopic manifestations.

Asthma↗

PubMed2003

Exogenous silicon (Si) increases antioxidant enzyme activity and reduces lipid peroxidation in roots of salt-stressed barley (Hordeum vulgare L.).

Two contrasting barley (Hordeum vulgare L.) cultivars, i.e. Kepin No.7 (salt sensitive) and Jian 4 (salt tolerant), were grown hydroponically to study the effect of exogenous silicon (Si) on time dependent changes of the activities of major antioxidant enzymes and of lipid peroxidation in roots under salt stress. Enzymes included: superoxide dismutase (SOD), peroxidase (POD), catalase (CAT) and glutathione reductase (GR). Three treatments with three replicates were investigated consisting of a control (basal nutrients with neither NaCl nor Si added), 120 mmol/L-1 NaCl, and 120 mmol/L-1 NaCl +1.0 mmol/L-1 Si. Plant roots were harvested 2, 4 and 6 days after treatment and assayed for activities of the antioxidant enzymes and the concentrations of reduced glutathione (GSH) and malondialdehyde (MDA), and electrolytic leakage percentage (ELP). The activities of SOD, POD and CAT in roots of salt-stressed plants were significantly stimulated at Day 2 compared to control plants, but considerably decreased at Day 4 and onward. GR activity in roots of salt-stressed plants remained unchanged at Day 2, but significantly decreased at Day 4 and onward. However, exogenous Si significantly enhanced these enzyme activities in roots of salt-stressed plants compared to Si-deprived salt treatments. This Si effect was time-dependent and became stronger as the experiments continued. The tendency of change in the activities of antioxidant enzymes and the concentration of GSH coincided with the concentration of MDA, the end product of lipid peroxidation, and the ELP. Higher activities of antioxidant enzymes, and higher concentration of GSH, but lower concentration of MDA and lower ELP were noted in cultivar Jian 4 compared to Kepin No. 7, implying genotypic differences with Jian 4 being less susceptible to stress-dependent membrane lipid peroxidation. The effects of Si-enhanced salt tolerance are discussed with respect to cell membrane integrity, stability and function in barley.

Antioxidants↗

PubMed2003

Molecular cytogenetic discrimination and reaction to wheat streak mosaic virus and the wheat curl mite in Zhong series of wheat--Thinopyrum intermedium partial amphiploids.

Thinopyrum intermedium (2n = 6x = 42, JJJsJsSS) is potentially a useful source of resistance to wheat streak mosaic virus (WSMV) and its vector, the wheat curl mite (WCM). Five partial amphiploids, namely Zhong 1, Zhong 2, Zhong 3, Zhong 4, and Zhong 5, derived from Triticum aestivum x Thinopyrum intermedium crosses produced in China, were screened for WSMV and WCM resistance. Zhong 1 and Zhong 2 had high levels of resistance to WSMV and WCM. The other three partial amphiploids, Zhong 3, 4, and 5, were resistant to WSMV, but were susceptible to WCM. Genomic in situ hybridization (GISH) using a genomic DNA probe from Pseudoroegneria strigosa (SS, 2n = 14) demonstrated that two partial amphiploids, Zhong 1 and Zhong 2, have almost the identical 10 Th. intermedium chromosomes, including four Js, four J, and two S genome chromosomes. Both of them carry two pairs of J and a pair of Js genome chromosomes and two different translocations that were not observed in the other three Zhong lines. The partial amphiploids Zhong 3, 4, and 5 have another type of basic genomic composition, which is similar to a reconstituted alien genome consisting of four S and four Js genome chromosomes of Th. intermedium (Zhong 5 has two Js chromosomes plus two Js-W translocations) with six translocated chromosomes between S and Js or J genomes. All three lines carry a specific S-S-Js translocated chromosome, which might confer resistance to barley yellow dwarf virus (BYDV-PAV). The present study identified a specific Js2 chromosome present in all five of the Zhong lines, confirming that a Js chromosome carries WSMV resistance. Resistance to WCM may be linked with J or Js chromosomes. The discovery of high levels of resistance to both WSMV and WCM in Zhong 1 and Zhong 2 offers a useful source of resistance to both the virus and its vector for wheat breeding programs.

Agropyron↗

PubMed2003

Molecular characterization of a wheat--Thinopyrum ponticum partial amphiploid and its derivatives for resistance to leaf rust.

Leaf rust (caused by Puccinia triticina Eriks.) occurs annually in most wheat-growing areas of the world. Thinopyrum ponticum (Podp.) Z.-W. Liu & R.-C. Wang has provided several leaf rust resistance genes to protect wheat from this fungal disease. Three chromosome substitution lines, Ji806, Ji807, and Ji859, and two chromosome addition lines, Ji791 and Ji924, with a winter growing habit were developed from crosses between wheat (Triticum aestivum L. em Thell.) and the wheat - Th. ponticum partial amphiploid line 693. These lines were resistant to leaf rust isolates from China. Sequence-tagged site (STS) analysis with the J09-STS marker, which is linked to the gene Lr24, revealed that the partial amphiploid line 693 and all of the substitution and addition lines carried gene Lr24. Genomic in situ hybridization (GISH) analysis was carried out on chromosome preparations using total genomic DNA from Pseudoroegneria strigosa (M. Bieb) A. Löve (St genome, 2n = 14) as a probe in the presence of total genomic DNA from T. aestivum 'Chinese Spring' wheat (ABD genomes, 2n = 42). The GISH analysis demonstrated that these lines had a pair of chromosomes displaying the typical pattern of a Js genome chromosome. This indicates that the chromosome that carries gene Lr24 belonged to the Js genome of Th. ponticum. In addition to 40 wheat chromosomes, eight Js and eight J genome chromosomes were also differentiated by GISH in the partial amphiploid line 693. Since most sources of Lr24 have a red grain color, the white-colored seeds in all of these substitution and addition lines, together with high protein content in some of the lines, make them very useful as a donor source for winter wheat breeding programs.

Chromosomes, Plant↗

PubMed2003

[Optimization of T7-based RNA amplification system for cDNA microarray].

cDNA microarrays are powerful parallel tools for gene expression profiling analysis, which help us to understand the molecular mechanism of diseases and to identify potential targets for therapeutic intervention. However, their broader application are hampered by the large amount of RNA required: up to 200 microg of total RNA or 5 microg of mRNA for one chip, making analysis of small samples difficult. In this work, combined with a template switching effect, the T7 RNA linear amplification procedure was optimized, providing multiple copies of anti-sense RNA with reduced sample inputs: no more than 3 microg total RNA for one chip. Using the same RNA sample in all cases, the anti-sense RNA labeling method was compared with standard total RNA and mRNA methods by two sets of self-comparison experiments. Furthermore, the three methods in profiling analysis were compared with the same pair RNA samples. All the results indicated that the fidelity, reproducibility, and reliability showed no significant difference with conventional total RNA or mRNA microarrays.

DNA-Directed RNA Polymerases↗

PubMed2003

Effects of transgene Oncostatin M on the development of retinal neuron in transgenic mice.

PURPOSE: Oncostatin M (OSM) is a cytokine released by macrophages and lymphocytes that can function as a growth regulator. A current study shows that leukemia inhibitory factor (LIF), a homologue of OSM, can prevent photoreceptor cell death when expressed in the lens of transgenic mice. We determined the efforts of lens-specific overexpression of OSM on the development of eye. METHODS: A truncated mouse OSM cDNA (-660 bp) was linked to the alpha A-crytallin promoter, and injected into single-cell embryos with microinjection. Then, transgenic mice were established. The mRNA expression of transgene OSM was detected by in situ hybridization. Immunohistochemistry was used to detect the expression of syntaxin, glial fibrillary acidic protein (GFAP), synaptophysin in the retinas of transgenic mice. RESULTS: At embryonic day (E 17.5), the expression of the syntaxin at the inner and mid portion of the retinas of transgenic mice was much higher than that of the retinas of non-transgenic mice. The expression of GFAP was detected in the retinas of transgenic mice, while no expression in non-transgenic normal FVB (FVB/N) mice was detected in this stage. At postnatal day one (P1), the expression of synaptophysin was detected in the retinas of transgenic mice, but there was no such expression in FVB/N mice. CONCLUSIONS: Lens-specific overexpression of OSM induces premature differentiation of amacrine cells, gial cells, and photoreceptors in vivo.

Animals↗

PubMed2003

[Validation of cDNA microarray technology].

cDNA microarray is a technological approach that has the potential to globally measure changes in mRNA expression levels. Self-comparison experiments with the same kind of tissue and differential expression experiments with the different kinds of tissue have been done to verify the reproducibility and the accuracy of this technique. The parameter of the reliability and the reproducibility of the microarray data were analyzed by correlation coefficient (R), coefficient of variation (CV) and false positive rate (FPR) etc. Meanwhile, the error resource also has been inspected. These results showed that generally the correlation coefficient of data from this cDNA microarray system was more than 0.9, the coefficient of variation was about 15%, and the false positive rate was below 3%. The result proves the accuracy of the cDNA microarray data. Consistence rate (CR) was advanced here as a new parameter to evaluate the reproducibility of two replicate experiments. It has some advantages over correlation coefficient and coefficient of variation. The influence of some important factors in the experiments, such as different concentration of spotted DNA, mRNA and total RNA, different batches of slides and different processes of labeling, have been investigated by comparing the results. It was shown that most of the false position produced by the experiment system could be reduced by replicate experiments.

Humans↗