Expression and role of anti-oxidative damage factors in the placenta of preterm infants with premature rupture of membranes. / æŠ—æ°§åŒ–æŸä¼¤å› å­åœ¨èƒŽè†œæ—©ç ´æœªè¶³æœˆå„¿èƒŽç›˜ä¸­çš„è¡¨è¾¾åŠä½œç”¨.

OBJECTIVES: To study the association of the anti-oxidative damage factors nuclear factor erythroid 2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), and NAD(P)H:quinone oxidoreductase-1 (NQO1) with preterm premature rupture of membranes (PPROM). METHODS: A prospective study was conducted. The neonates who were hospitalized in Yanbian Hospital from 2019 to 2020 were enrolled as subjects, among whom there were 30 infants with PPROM, 32 infants with term premature rupture of membranes (TPROM), and 35 full-term infants without premature rupture of membranes (PROM). Hematoxylin and eosin staining was used to observe the inflammatory changes of placental tissue. Immunohistochemical staining was used to measure the expression of Nrf2, HO-1, and NQO1 in placental tissue. Western blot was used to measure the protein expression levels of Nrf2, HO-1, and NQO1 in placental tissue. RESULTS: Compared with the PPROM group, the TPROM group and the non-PROM full-term group had significantly higher positive expression rates and relative protein expression levels of Nrf2, HO-1, and NQO1 in placental tissue (P<0.05). There were no significant differences in the positive expression rates and relative protein expression levels of Nrf2, HO-1, and NQO1 in placental tissue between the TPROM and non-PROM full-term groups (P>0.05). CONCLUSIONS: The low expression levels of Nrf2, HO-1, and NQO1 in placental tissue may be associated with PPROM, suggesting that anti-oxidative damage is one of the directions to prevent PPROM.

Effect of the phthalates exposure on sex steroid hormones in the US population.

BACKGROUND: New alternative phthalates have been increasingly substituted for certain phthalates in some consumer products due to safety concerns. However, research on the steroidal effect of exposure to the newer replacement phthalates in the general adult population is lacking. OBJECTIVES: This study aimed to examine the associations of exposure to the older generation and newer replacement phthalates with sex hormone levels in the U.S. general population. METHODS: The current cross-sectional study was based on the National Health and Nutrition Examination Survey (NHANES) 2015-2016. Sixteen urinary phthalates metabolites and three serum sex hormones were measured in 1768 adults. Gender-specific associations between urinary phthalate concentrations and sex hormones were estimated by using adjusted multiple linear regression. Logistic regression was performed to calculate the risk of phthalates exposure on hormones dysfunction. RESULTS: Most phthalates metabolites concentrations were lower than 50 ng/mL. MEP, MBP, MiBP, MECPP, MCOP, MEHHP, MEOHP were higher than others, suggesting that new alternative DEP, DBP, and DiNP were exposed at high levels in daily life while DINCH was at a low level. Phthalates exposure was associated with decreased testosterone levels and increased estradiol and SHBG in total samples. Testosterone level was negatively associated with MnBP (ß: -0.05, 95% CI: -0.09, 0), MEOHP (ß:-0.05, 95% CI:-0.09,-0.01), MEHHP (ß:-0.04, 95% CI:-0.08,0), MECPP (ß:-0.07, 95% CI:-0.11,-0.03), MEP (ß: -0.03, 95% CI: -0.06, 0), MiBP (ß: -0.05, 95% CI: -0.10, -0.01) in males; ln-transformed estradiol were increased by 0.18 pg/mL (95% CI: 0.05,0.31), 0.15 pg/mL (95% CI: 0.01,0.29) with each 1 ln-concentration increase in MEHP and MNP, respectively, in females. CONCLUSIONS: Our results suggest that phthalates exposure may disturb the hormone homeostasis in adults. The safe alternative should be used with caution in industrial production in the future and the need for further research into the safety of the new alternative replacements is necessary.

Development of an UPLC-MS/MS Method for the Quantitative Analysis of Upadacitinib in Beagle Dog Plasma and Pharmacokinetics Study.

BACKGROUND: Upadacitinib, a novel selective Janus kinase 1 (JAK1) inhibitor, has been recently approved by the US FDA for the treatment of adult patients with moderately to severely active rheumatoid arthritis (RA). An ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method for the quantitative analysis of upadacitinib in beagle dog plasma was developed and validated. METHODS: Upadacitinib and fedratinib (internal standard, IS) were extracted with ethyl acetate under alkaline condition and then separated and detected. The chromatographic column was Waters Acquity UPLC BEH C18 column (2.1 mm × 50 mm, 1.7 µm), the mobile phase was acetonitrile and 0.1% formic acid in water with gradient elution procedure, and the flow rate was 0.40 mL/min. Under the positive ion mode, upadacitinib and IS were monitored by multiple reaction monitoring (MRM) as the following mass transition pairs: m/z 447.00 → 361.94 for upadacitinib and m/z 529.82 → 141.01 for IS. RESULTS: In the concentration range of 1-500 ng/mL, upadacitinib had good linearity, and the lower limit of quantification (LLOQ) was 1 ng/mL. The RSD of the intra- and inter-day precision was less than 10.03%, and the RE of accuracy was -3.79% to 2.58%. The extraction recovery of upadacitinib was more than 80%, the matrix effect was around 100%, and upadacitinib was found to be stable. CONCLUSION: The novel optimized UPLC-MS/MS assay was an effective tool for the determination of upadacitinib and had been successfully applied to the pharmacokinetic study of upadacitinib in beagle dogs, and this method would also be used to study DDIs.

Characterization and exposure assessment of household fine particulate matter pollution in China.

Household fine particulate matter (PM2.5 ) pollution greatly impacts residents' health. To explore the current national situation of household PM2.5 pollution in China, a study was conducted based on literature published from 1998 to 2018. After extracting data from the literature in conformity with the requirements, the nationwide household-weighted mean concentration of household PM2.5 (HPL) was calculated. Subgroup analyses of spatial, geographic, and temporal differences were also done. The estimated overall HPL in China was 132.2 ± 117.7 µg/m3 . HPL in the rural area (164.3 ± 104.5 µg/m3 ) was higher than that in the urban area (123.9 ± 122.3 µg/m3 ). For HPLs of indoor sampling sites, the kitchen was the highest, followed by the bedroom and living room. There were significant differences of geographic distributions. The HPLs in the South were higher than the North in four seasons. The inhaled dose of household PM2.5 among school-age children differed from provinces with the highest dose up to 5.9 µg/(kg·d). Countermeasures should be carried out to reduce indoor pollution and safeguard health urgently.

Source, Characterization of Indoor Dust PAHs and the Health Risk on Chinese Children.

Polycyclic aromatic hydrocarbons (PAHs) in indoor dust are one of the common exposure sources for children worldwide. The aim of this study is to explore PAHs pollution status in indoor dust and estimate health risk on Chinese children with big data. Weighted average concentration was used to analyze source and characterization of PAHs in indoor dust based on peer-reviewed literature. According to specific inclusion criteria, 17 studies were included finally to analyze weighted average concentration. The national average concentration of Σ16PAHs was approximately 25.696 µg/g. The highest concentration of Σ16PAHs was in Shanxi (2111.667 µg/g), and the lowest was in Hong Kong (1.505 µg/g). The concentrations in Shanxi and Guangdong were higher than national level and the over standard rate was 18.18%. The concentrations of individual PAHs varied greatly across the country, and Flu in Shanxi was the highest (189.400 µg/g). The sources of PAHs varied in different regions and combustion processes played a leading role. PAHs exposure through ingestion and dermal contact was more carcinogenic than inhalation. The incremental lifetime cancer risk model indicated that children lived in Shanxi were found in the highest health risk coupled with the highest BaPE concentration (54.074 µg/g). Although PAHs concentrations of indoor dust showed a downward trend from 2005 to 2018, indoor environmental sanitation should be improved with multidisciplinary efforts. Health standard should be possibly established to minimize children exposure to PAHs in indoor dust in China.

[Effect and Mechanism of Foliar Application Nano-MnO2 on Cadmium Enrichment of Rice].

Cadmium (Cd) is easily enriched in rice, resulting in an excessive Cd content in the grain, which seriously threatens human health. Manganese (Mn) is an essential element of plants. In a field experiment on Cd-contaminated acid soils, we investigated the effectiveness and mechanism of Mn in minimizing Cd accumulation in rice via foliar spraying using 0.1%, 0.3%, and 0.5% nano-MnO2 solutions at an early stage of heading. Compared with a control treatment, foliar spraying effectively reduced the Cd content of rice leaves, husks, and brown rice; increased the Mn content of all rice organs; but had no effect on rice yield. Foliar application of nano-MnO2 alleviated the plant stress induced by Cd by improving leaf photosynthesis, inhibiting lipid peroxidation, and increasing the content of the oxidative stress protein kinase. In addition, foliar application of nano-MnO2 limited the absorption of Cd by roots by increasing the content of iron-manganese plaque on the surfaces of rice roots and strengthening its adsorption/co-precipitation of Cd. Therefore, foliar application of nano-MnO2 during the early stages of rice heading is an effective measure to increase the Mn content and reduce the Cd content of brown rice.

Predictive Value of Pin1 in Cervical Low-Grade Squamous Intraepithelial Lesions and Inhibition of Pin1 Exerts Potent Anticancer Activity against Human Cervical Cancer.

Many oncogenes are involved in the progression from low-grade squamous intraepithelial lesions (LSILs) to high-grade squamous intraepithelial lesions (HSILs); which greatly increases the risk of cervical cancer (CC). Thus, a reliable biomarker for risk classification of LSILs is urgently needed. The prolyl isomerase Pin1 is overexpressed in many cancers and contributes significantly to tumour initiation and progression. Therefore, it is important to assess the effects of cancer therapies that target Pin1. In our study, we demonstrated that Pin1 may serve as a biomarker for LSIL disease progression and may constitute a novel therapeutic target for CC. We used a the novel Pin1 inhibitor KPT-6566, which is able to covalently bind to Pin1 and selectively target it for degradation. The results of our investigation revealed that the downregulation of Pin1 by shRNA or KPT-6566 inhibited the growth of human cervical cancer cells (CCCs). We also discovered that the use of KPT-6566 is a novel approach to enhance the therapeutic efficacy of cisplatin (DDP) against CCCs in vitro and in vivo. We showed that KPT-6566-mediated inhibition of Pin1 blocked multiple cancer-driving pathways simultaneously in CCCs. Furthermore, targeted Pin1 treatment suppressed the metastasis and invasion of human CCCs, and downregulation of Pin1 reversed the epithelial-mesenchymal transition (EMT) of CCCs via the c-Jun/slug pathway. Collectively, we showed that Pin1 may be a marker for the risk of progression to HSIL and that inhibition of Pin1 has anticancer effects against CC.

[Variations in topsoil carbon and nitrogen contents of five temperate plantations in Northeast China.] / 东北5种温带人工林表层土壤碳氮含量的分异.

Afforestation is an effective way for carbon (C) sequestration, which also profoundly influences soil nitrogen (N) dynamics in the forest ecosystem. The impacts of tree species on soil C and N budgets and the underlying mechanism remain uncertain. In this study, we used a common garden experiment and measured the soil organic C (Csoil) and total N contents (Nsoil) of the topsoil (0-10 cm) and related vegetative and soil microbial properties in 2007 and 2015 (3 and 11 years after afforestation), respectively. Our aim was to explore the effects of five major tree species (i.e., Fraxinus mandshurica, Juglans mandshurica, Betula platyphylla, Larix gmelinii and Pinus sylvestris var. mongolica) in the temperate forests in Northeast China on soil C and N contents and their dri-vers. The results showed that both Csoil and Nsoil of the five stands decreased as the stand ages increased, the change rates of which were significantly correlated with each other. The rate of change in Csoil(2.6%·a-1-4.8%·a-1) was significantly greater than that in Nsoil(0.8%·a-1-2.8%·a-1). The decrements of the Csoil and Nsoil for the broadleaved stands were significantly lower than those of the coniferous stands. The tree-species traits and microbial properties together explained 68.5% and 90.9% of the variability of the change rates of Csoil and Nsoil, respectively. The change rates of Csoil and Nsoil decreased with the increases in leaf litter C/N and microbial biomass C/N, but increased with the increases of fine root biomass, microbial biomass C, and the ratio of the C-acquisition to the N-acquisition enzyme activity. Additionally, the change rate of Nsoil decreased with the increases of the leaf litter production and the microbial metabolic activity. Our findings indicated that C and N contents in the topsoil of these temperate plantations decreased significantly 11 years after afforestation, while the different change rates mainly resulted from different properties of tree species and soil microbes.

Exendin-4 Exacerbates Burn-Induced Morbidity in Mice by Activation of the Sympathetic Nervous System.

BACKGROUND: Although glucagon-like peptide 1- (GLP-1-) based therapy of hyperglycemia in burn injury has shown great potential in clinical trials, its safety is seldom evaluated. We hypothesize that exendin-4, a GLP-1 analogue, might affect the immune response via the activation of the sympathetic nervous system in burn injury. METHODS: Male Balb/c mice were subjected to sham or thermal injury of 15% total body surface area. Exendin-4 on T cell function in vitro was examined in cultured splenocytes in the presence of ß-adrenoceptor antagonist propranolol (1 nmol/L) or GLP-1R antagonist exendin (9-39) (1 µmol/L), whereas its in vivo effect was determined by i.p. injection of exendin-4 (2.4 nmol/kg) in mice. To further elucidate the sympathetic mechanism, propranolol (30 mg/kg) or vehicle was applied 30 min prior to injury. RESULTS: Although the exacerbated burn-induced mortality by exendin-4 was worsened by propranolol pretreatment, the inhibition of T cell proliferation by exendin-4 in vitro could be restored by propranolol instead of exendin (9-39). However, a Th2 switch by exendin-4 in vitro could only be reversed by exendin (9-39). Likewise, the inhibition of splenic T cell function and NFAT activity by exendin-4 in vivo was restored by propranolol. By contrast, the increased splenic NF-κB translocation by exendin-4 in vivo was potentiated by propranolol in sham mice but suppressed in burn mice. Accordingly, propranolol abrogated the heightened inflammatory response in the lung and the accelerated organ injuries by exendin-4 in burn mice. On the contrary, a Th2 switch and higher serum levels of inflammatory mediators by exendin-4 were potentiated by propranolol in burn mice. Lastly, exendin-4 raised serum stress hormones which could be remarkably augmented by propranolol. CONCLUSIONS: Exendin-4 suppresses T cell function and promotes organ inflammation through the activation of the sympathetic nervous system, while elicits Th2 switch via GLP-1R in burn injury.

Gene expression profiling of flag leaves at the booting stage in the japonica hybrid rice Huayou14 and its parental lines by microarray.

Gene expression profiling using microarray has contributed significantly to heterosis studies. Using the Affymetrix rice genome array, we investigated gene expression profiles in the flag leaves of the japonica hybrid rice Huayou14 and its parental cultivars Shen9A and Fan14 at the booting stage. A total of 2057 genes differentially expressed (fold change ≥2 or ≤0.5) between Huayou14 and its parents were identified. Functional classification of the differentially expressed genes by Gene Ontology (GO) analysis indicated the differentially expressed genes were significantly enriched in photosynthesis-related cellular component categories (e.g. photosystem Ⅰ, chloroplast membrane and chloroplast envelope), and biological process categories (e.g. chlorophyll catabolic, chlorophyll biosynthetic and carotenoid biosynthetic processes). These results suggest that the changes in the photosynthetic ability of the japonica hybrid rice Huayou14 may be related to heterosis. Metabolic pathway analysis indicated that differentially expressed genes were significantly enriched in photosynthesis-antenna proteins and starch and sucrose metabolic pathways, instead of photosynthesis and carbon fixation pathways as reported previously. These results suggest that different genes or metabolic pathways might contribute to the heterosis of different hybrid combinations.

An outbreak following importation of wild poliovirus in Xinjiang Uyghur Autonomous Region, China, 2011.

BACKGROUND: After more than 10 years without a case of wild poliovirus (WPV) in China, an outbreak occurred in 2011 in Xinjiang Uyghur Autonomous Region. METHODS: Acute flaccid paralysis (AFP) case surveillance was strengthened with epidemiological investigations and specimen collection and serological surveys were conducted among hospitalized patients. RESULTS: There were 21 WPV cases and 23 clinical compatible polio cases reported. WPV was isolated from 14 contacts of AFP cases and 13 in the healthy population. Incidence of WPV and clinical compatible polio cases were both highest among children <1 years, however, 24/44 (54.5%) polio cases were reported among adults aged 15-39 years. CONCLUSIONS: High coverage of routine immunization should be maintained among children until WPV transmission is globally eradicated. Expansion of AFP case surveillance and use of serologic surveys to estimate population immunity should be conducted rapidly to guide preparedness and response planning for future WPV outbreaks.

Enhanced surveillance of acute flaccid paralysis following importation of wild poliovirus in Xinjiang Uygur Autonomous Region, China.

BACKGROUND: After being polio free for more than 10 years, an outbreak occurred in China in 2011 in Xinjiang Uygur Autonomous Region (Xinjiang) following the importation of wild poliovirus (WPV) originating from neighboring Pakistan. METHODS: To strengthen acute flaccid paralysis (AFP) surveillance in Xinjiang, "zero case daily reporting" and retrospective searching of AFP cases were initiated after the confirmation of the WPV outbreak. To pinpoint all the polio cases in time, AFP surveillance system was expanded to include persons of all ages in the entire population in Xinjiang. RESULTS: Totally, 578 AFP cases were reported in 2011 in Xinjiang, including 21 WPV cases, 23 clinical compatible polio cases and 534 non-polio AFP cases. Of the 44 polio cases, 27 (61.4%) cases were reported among adults aged 15-53 years. Strengthening AFP surveillance resulted in an increase in the number of non-polio AFP cases in 2011 (148 children < 15 years) compared with 76 cases < 15 years in 2010. The AFP surveillance system in Xinjiang was sensitive enough to detect polio cases, with the AFP incidence of 3.28/100,000 among children < 15 years of age. CONCLUSIONS: Incorporating adult cases into the AFP surveillance system is of potential value to understand the overall characteristics of the epidemic and to guide emergency responses, especially in countries facing WPV outbreak following long-term polio free status. The AFP surveillance system in Xinjiang was satisfactory despite limitations in biological sample collection.

Identification and control of a poliomyelitis outbreak in Xinjiang, China.

BACKGROUND: The last case of infection with wild-type poliovirus indigenous to China was reported in 1994, and China was certified as a poliomyelitis-free region in 2000. In 2011, an outbreak of infection with imported wild-type poliovirus occurred in the province of Xinjiang. METHODS: We conducted an investigation to guide the response to the outbreak, performed sequence analysis of the poliovirus type 1 capsid protein VP1 to determine the source, and carried out serologic and coverage surveys to assess the risk of viral propagation. Surveillance for acute flaccid paralysis was intensified to enhance case ascertainment. RESULTS: Between July 3 and October 9, 2011, investigators identified 21 cases of infection with wild-type poliovirus and 23 clinically compatible cases in southern Xinjiang. Wild-type poliovirus type 1 was isolated from 14 of 673 contacts of patients with acute flaccid paralysis (2.1%) and from 13 of 491 healthy persons who were not in contact with affected persons (2.6%). Sequence analysis implicated an imported wild-type poliovirus that originated in Pakistan as the cause of the outbreak. A public health emergency was declared in Xinjiang after the outbreak was confirmed. Surveillance for acute flaccid paralysis was enhanced, with daily reporting from all public and private hospitals. Five rounds of vaccination with live, attenuated oral poliovirus vaccine (OPV) were conducted among children and adults, and 43 million doses of OPV were administered. Trivalent OPV was used in three rounds, and monovalent OPV type 1 was used in two rounds. The outbreak was stopped 1.5 months after laboratory confirmation of the index case. CONCLUSIONS: The 2011 outbreak in China showed that poliomyelitis-free countries remain at risk for outbreaks while the poliovirus circulates anywhere in the world. Global eradication of poliomyelitis will benefit all countries, even those that are currently free of poliomyelitis.

Overexpression of ACL1 (abaxially curled leaf 1) increased Bulliform cells and induced Abaxial curling of leaf blades in rice.

Understanding the genetic mechanism underlying rice leaf-shape development is crucial for optimizing rice configuration and achieving high yields; however, little is known about leaf abaxial curling. We isolated a rice transferred DNA (T-DNA) insertion mutant, BY240, which exhibited an abaxial leaf curling phenotype that co-segregated with the inserted T-DNA. The T-DNA was inserted in the promoter of a novel gene, ACL1 (Abaxially Curled Leaf 1), and led to overexpression of this gene in BY240. Overexpression of ACL1 in wild-type rice also resulted in abaxial leaf curling. ACL1 encodes a protein of 116 amino acids with no known conserved functional domains. Overexpression of ACL2, the only homolog of ACL1 in rice, also induced abaxial leaf curling. RT-PCR analysis revealed high expressions of ACLs in leaf sheaths and leaf blades, suggesting a role for these genes in leaf development. In situ hybridization revealed non-tissue-specific expression of the ACLs in the shoot apical meristem, leaf primordium, and young leaf. Histological analysis showed increased number and exaggeration of bulliform cells and expansion of epidermal cells in the leaves of BY240, which caused developmental discoordination of the abaxial and adaxial sides, resulting in abaxially curled leaves. These results revealed an important mechanism in rice leaf development and provided the genetic basis for agricultural improvement.

[Transpositional behaviour analysis of Ds element from different insertion sites on chromosome 4 in rice].

Transgenic plants with Ds element distributed over different loci on chromosome 4 (Fig. 1) and the homozygous transformants with Ac transposase gene were established through Agrobacterium-mediated approach. In this study, the plants carrying Ds element from different loci were crossed with the plant carrying Ac transposase individually. The plants of F(1) generation carrying both Ds element and Ac transposase were used to produce the F(2) populations (Table 1). Analysis of the F(2) generation by the PCR method revealed that the excision frequencies of Ds element were higher in the telomeric region of chromosome 4 than in the centromeric region (Fig. 4). These results showed that the insertion site of Ds element has strong effect on its excision frequency. We suggest that the special construct of chromosome near the insertion site of Ds element is related to the excision frequency of the Ds element.

[Establishment of promoter trapping system mediated by activator/dissociation (beta-glucuronidase) construction in rice].

The coding region of Bar gene, the left border of Ds element, the coding region of GUS gene, the transposase of Ac element, the right border of Ds element and the promoter of Ubi gene were inserted into the T-DNA region of vector pCAMBIA1300 in turn to construct plasmid p13B. The orientations of the ubiquitons' promoter, Ac transposase and Bar are identical but opposite to that of the GUS gene (Fig.1). The plasmid p13B was then introduced into the calli of Oryza sativa subsp. japonica cv. Zhonghua 11 by Agrobacterium tumefaciens-mediated transforming to trap genes in rice. Eighteen independent transgenic lines were obtained and propagated. T(2) generations of 18 independent transgenic lines were screening by herbicide (Basta) (Fig.2) and the herbicide-resistant plants obtained were analyzed by PCR (Fig.3). Ds element transposed in an inheritable manner was found in 37 plants, in which 5 plants showed GUS activity (Fig.4).

[Determination of amylose content in transgenic progenies containing an antisense waxy gene and identifying of pure lines in indica rice].

Through genetic transformation mediated by Agrobacterium tumefaciens, an antisense waxy gene was introduced into Longtefu B line, a male sterile maintainer line in indica (Oryza sativa L.). Thirty transgenic plants showed integration of antisense waxy gene into the genome as determined by PCR assay, and twenty eight were confirmed by Southern blotting. T1 seeds from twenty one transgenic plants showed a marked decrease of amylose content, ranging from 3%-13% less than control, and seeds from some transgenic lines exhibited typical waxy phenotype. Six transgenic lines were selected to examine the amylose content in different generations. In the T4 generation, two homozygous lines, Long 3-1-1-1 and Long 5-8-2-1, were selected, with amylose content of 15.9% and 8.4% respectively. The amylose content in these lines is in consistent with the decrease of the accumulation of Wx protein as determined by SDS-PAGE electrophoresis. Cross and subsequent backcross of Long 3-1-1-1 and Long 5-8-2-1 with the Longtefu mare sterile line were performed to determine the changes of amylose content in F1 and B1F1 seeds. The results showed an average amlyose content of 21.4% in F1 seeds with Long 3-1-1-1 as a parent, while only 13.6% with Long 5-8-2-1 as a parent. In addition, the average amylose contents in B1F1 seeds were 17.1% and 9.3% respectively. Our results indicated that during the fertility transfer in the male sterile line, stable transgenic lines with medium or low contents of amylose had direct effects on amylose content in F1 seeds.

[Rapidly obtaining the markerless transgenic rice with reduced amylose content by co-transformation and anther culture].

The plasmid p13W8 carrying antisense fragment of waxy gene and plasmid pCAMBIA1300 containing hpt gene were introduced into rice by Agrobacterium tumefaciens-mediated co-transformation, and 86 transgenic plants were obtained, 32 of them showed positive bands for antisense waxy gene by PCR analysis, the waxy-positive plant frequency is 37.2%. The segregation of antisense fragment of waxy gene and hpt gene was observed by PCR using hpt gene primers and waxy gene primers respectively in 29 T(1) population. One hundred and eighty-three plants containing only the antisense fragment of waxy gene were identified in 1 264 T(1) plants, the waxy-positive plant frequency is 14.4% (Table 1). The amylose content of seeds derived from transgenic plants with only the antisense fragment of waxy gene were determined, varying degrees of reduction in amylose content were found in some plants (Table 2). Four T(1) plants with reduced amylose content were selected through anther culture. Thirty-four anther culture plants seed normally, 23 of them were shown to contain only the antisense fragment of waxy gene (Table 3) by PCR analysis, and the amylose content was reduced to 5%-12% (Table 4). It took only one and half years to obtain the stably inherited markerless transgenic rice with reduced amylose content by co-transformation and anther culture technique.

[Genetic analysis of a rolled-leaf mutant in rice population of T-DNA insertion].

A rolled-leaf mutant was obtained in a T-DNA(containing bar gene and Ds element) insertion population, which consist of transgenic japonica rice Zhonghua 11 mediated by Agrobacterium tumefaciens. Through self-hybridization of three generations, one of trait-purified mutants (R1-A2) was obtained and used as parent to cross with variety Zhonghua 11. The leaves of 36 F1 plants investigated were rolled and resistant to herbicide Basta. Among 852 F2 plants, the segregation ratio of rolled leaves to normal leaves(645:207) was consistent with 3:1. All rolled-leaf plants were resistant to herbicide Basta, and all normal leaf plants were sensitive to herbicide Basta. These results showed that the trait of rolled-leaf is co-segregated with Basta resistance. The total DNA of 45 rolled-leaf plants and 30 normal leaf plants in F2 population were amplified to test the presence of T-DNA by Ds primers. The results showed that the positive band were amplified in all rolled-leaf plants, but not in every normal leaf plant. In F1B1 progenies, all plants which derived from backcross parent R1-A2 were rolled leaves; while variety Zhonghua 11 was used as backcross parent, the segregation ratio of rolled-leaf to normal leaf was consistent with 1:1. Taking these data together, it indicated that the rolled-leaf mutant was co-segregation with T-DNA and controlled by single dominant gene.