Genetic Analysis and Molecular Mapping of Light-Sensitive Red-Root Mutant in Rice

The light-sensitive red-root mutant, designated as HG1, was newly observed from an indica rice variety, Nankinkodo, when seedlings were grown with roots exposed to natural light. The root color of the mutant began to turn slight-red when the roots were exposed to the light at the intensity of 29 ）Jmol/（m^2·s）, then turned dark-red at the light intensity of 180 pmol/（m^2·s）, suggesting that the root color of the mutant was evidently sensitive to light. Furthermore, genetic analysis showed that the character of light-sensitive red-root of the HG1 mutant was controlled by a single dominant gene, tentatively designated as Lsr. With simple sequence repeat markers, Lsrgene was located between the markers RM252 and RM303 on chromosome 4 with the genetic distances of 9.8 cM and 6.4 cM, respectively. These results could be useful for fine mapping and cloning of Lsrgene in rice.

Genetic Analysis and Primary Mapping of pms4, a Photoperiod-Sensitive Genic Male Sterility Gene in Rice (Oryza sativa)

To understand the genetic characteristics of a new photoperiod-sensitive genic male sterile line Mian 9S, some reciprocal crosses were made between Mian 9S and six indica rice materials, Yangdao 6, Luhui 602, Shuihui 527, Mianhui 725, Fuhui 838 and Yixiang 1B. Genetic analysis results suggested that the photoperiod-sensitive genic male sterility （PGMS） of Mian 9S was controlled by a single recessive nuclear gene. Thus, the F2 population derived from the cross of Yangdao 6/Mian 9S was used to map the PGMS gene in Mian 9S. By using SSR markers, the PGMS gene of Mian 9S was mapped on one side of the markers, RM6659 and RM1305, on rice chromosome 4, with the genetic distances of 3.0 cM and 3.5 cM, respectively. The gene was a novel PGMS gene and designated tentatively as pms4. In addition, the application of the pms4 gene was discussed.

Identification of quantitative trait loci and candidate genes associated with ABA sensitivity in common wild rice (Oryza rufipogon Griff,)

Abscisic acid （ABA）, as one of the foremost signaling molecules in plants, is an important hormone which plays versatile functions in regulating developmental process and adaptive stress process. A set of introgression lines were previously generated via a backcrossing program using an elite indica cultivar rice Teqing （O. sativa L.） as recipient and an accession of Yuanjiang common wild rice （O. rufipogon Griff.） as donor. In this study, the previously developed introgression lines were evaluated for ABA sensitivity. Here we reported that a total of 14 quantitative trait loci （QTLs） associated with ABA sensitivity were identified. An ABA sensitive introgression line, YIL53, was identified and characterized. Physiological characterization, including chlorophyll content, malondialdehyde content, soluble sugar content, and stomata movement, demonstrated that YIL53 exhibited the characteristics associated with ABA sensitivity. Genotypic analysis revealed that YIL53 harbored one QTL related to ABA sensitivity, qASS1-2, which was located on chromosome 1 within one introgressed segment derived from the Yuanjiang common wild rice. Furthermore, the qASS1-2 was finally narrowed down to a 441-kb region between simple sequence repeats （SSR） marker RM212 and single nucleotide polymorphism （SNP） marker M3 using the segregation population derived from the cross between Teqing and YIL53, and three candidate genes associated with ABA sensitivity were identified using a strategy combined gene expression analysis with QTL mapping. Identification of the QTLs related to ABA sensitivity and characterization of the ABA sensitive line YIL53 would provide a helpful basis for isolating novel genes related to ABA sensitivity.

Progresses and Strategies of Gene Mapping and Isolating of Photo(thermo) sensitive Genic Male Sterile Rice

Considering the research on classical genetics of photoperiod(therm) sensitive genic male sterile rice, it is important to select the sterile lines and their segregating population controlled by one pair of gene in mapping and isolating sterile genes. It is discussed the advantages, disadvantages and the reasons leading to various mapping results of chromosome location of sterile genes through gene marker, isozyme marker and DNA marker techniques. In comparison to isolation of photo(thermo) sensitive sterile genes via various plant gene cloning techniques, it was concluded that map based cloning was acceptable, but it is still difficult to locate the loci of sterile genes within 1cM. On the other hand, “sensitivity to environment”, an important characteristic of sterile lines can be fully utilized by DD PCR and (or) RDA techniques. Therefore, these two techniques were considered as the effective ways to isolate sterile genes.

Genetic Analysis and Mapping of a Thermo-sensitive White Stripe-Leaf Mutant at Seedling Stage in Rice(Oryza sativa)

A thermo-sensitive white stripe-leaf mutant （tws） was selected from the M2 progeny of a japonica variety, Jiahua 1, treated by ^60 Co γ-radiation. In comparison with the wild type parent, the mutant displayed a phenotype of white stripe on the 3rd and 4th leaves, but began to turn normal green on the 5th leaf when grown at low temperatures （20℃ and 24℃）. Furthermore, the content of total chlorophyll showed an obvious decrease in the leaves with white stripe. These results suggest that the expression of the mutant trait was thermo-sensitive and correlated with the leaf age of seedlings. The genetic analysis indicated that the mutant trait was controlled by a single recessive nuclear gene, designated as tws. In addition, by using SSR markers and an F2 segregating population derived from the cross between the tws mutant and 9311, tws was mapped between the markers MM3907 and MM3928 with a physical distance of 86 kb on dce chromosome 4.

Studies on the Photoperiod Sensitive Characters of Male Fertility Alteration of Peiai64S' Main Male Genic Sterile Gene

Peiai64S, an indica male sterile rice with a male fertility alteration under different environments, is selected from the offspring of indica rice crossed with Nongken58S. Nongken58S, a japonica pho-toperiod sensitive genie male sterile rice (PGMS), deriving from a natural mutant plant individual of normal japonica rice variety, Nongken58, is used as a male sterile gene donor of Peiai64S. But Peiai64S is not a typical PGMS rice, the male fertility is sensitive to temperature just as thermo-sensitive genie male sterile rice (TGMS). We have selected typical PGMS plants in F2 population of Peiai64S× Nongken58, whose ratio of fertile plants to sterile plants is nearly 3:1. The sterility inheritance conformed to one pair of gene segregation model. The result indicates the main male sterile gene in Peiai64S is not other than the PGMS gene, and comes from Nongken58S. The genetic background affects effective expression of the PGMS gene. This suggests that we ought to focus on optimizing the genetic background of the PGMS gene in PGMS rice breeding, and select an ideal genetic background as a transgenic background in molecular breeding.

Effects of Mitofusin-2 Gene on Cell Proliferation and Chemotherapy Sensitivity of MCF-7

In order to evaluate the effect of mitofusin-2 gene （mfn2） on proliferation and chemotherapy sensitivity of human breast carcinoma cell line MCF-7 in vitro, pEGFPmfn2 plasmid carrying full length of mitofusin-2 gene was transfected, by using sofast, into MCF-7 cells. Mitofusin-2 gene expression in MCF-7 cells transfected by sofast after 48 h was detected by PCR and Western blotting, and the stable expression of GFP protein in MCF-7 cells by Western blot analysis. The proliferation of MCF-7 cells was assayed by MTT and cell counting. By using PI method, the effects of mfn2 on the cell cycle distribution of MCF-7 were measured. Annexin-Ⅴ/PI double labeling method was employed to detect the changes in apoptosis induced by chemotherapeutics before and after transfection. The results showed that the MCF-7 cells transfected with mfn2 gene could stably and highly express GFP protein. MTT assay revealed that after transfection of mfn2 cDNA, the proliferation of MCF-7 cells was significantly inhibited. DNA histogram showed that cells arrested in S phase, and the percentage of S phase cells was 42.7, 17.2 and 19.6 in mfn2 cDNA transfection group, blank plasmid transfection group and blank control group, respectively （P〈0.05）. The apoptosis ratio of the cells transfected with mfn2 gene was increased from 3.56% to 15.95%, that of the cells treated with camptothecin （CAMP） followed by mfn2 gene transfection was 69.6%, and that in blank plasmid transfection group and blank control group was 31.0% and 23.4% respectively （P〈0.05）. It was suggested that transfection of mfn2 gene could significantly inhibit the proliferation of MCF-7 cells and promote their sensitivity to CAMP with a synergic effect.

Studies on the Heredity of the Traits Related to the Photoperiod-Sensitive Phenomenon Among the Temperate ×Tropical Crosses in Maize

The genetic study of characters concerning photoperiod sensitivity of the growth and development stages, plant and ear height was carried out from 5 temperate tropical combinations by using P1 , P2, F1, F2, BC1, BC2 generations mean analysis. The results showed that the square sums of additive, of dominance and of epistatic gene effects contributed to total genetic variance were 11. 3 -53. 6%, 36.0 -78. 6% and 5.5 -27.0% respectively for five traits of the insensitive×insensitive combination to photoperiod, which suggested dominance gene effect was the most important among various gene effects. In insensitive X sensitive combination to photoperiod, the square sums of additive gene effect with 66. 9 - 84. 5 %, of dominance gene effect with 11. 6 30. 7% and of epistatic gene effect with 1.4-5. 2% were found for five tested traits, which showed additive gene effect held a dominant position, and dominance and epistatic gene effects were reduced compared with insensitive×insensitive combination. The differences existed among different insensitive×sensitive combinations. The proportion of the genetic component among the insensitive×sensitive crossbreed offspring in the temperate zone had a certain dosage effect on fading photoperiod sensitivity reaction. The sensitivity of BC1 , with 25 % of the tropical genetic component, is extremely weak, but with high application value.

Isolation and Analysis of the Promoter of OsRacD from PhotoperiodSensitive Genic Male Sterile Rice

By using OsRacD cDNA as probe to screen the genomic library of photoperiod sensitive genic male sterile rice line Nongken 58S, a positive clone containing 2 kb promoter and 396 bp coding region of OsRacD was obtained. Compared with the promoter of OsRacD cloned by reverse PCR from normal rice variety Nongken 58 （Nongken 58N）, the homology was 99.8%, and the different nucleotides were outside the predicted response elements in promoter, suggesting that the fertility between rice varieties Nongken 58S and Nongken 58N under the long-day conditions was not attributed to the difference in the structure of OsRacD upstream regulation sequences, but to the developmental regulation of gene differential expression.

Fertility Expression of TGMS-Genes in the Backgrounds of indica CMS-lines,B-lines and R-lines of Hybrid Rice

The generation fertility of 51 F1, 19 F2 and 6 BC1 between 3 thermo-sensitive genic male sterile lines （TGMS-lines） Pei＇ai 64S, 6311S and 360S, and the three lines of hybrid rice including 7 indica cytoplasmic male sterile lines （CMS-lines） and their corresponding maintainer lines （B-lines） and 3 indica restorer lines （R-lines） were investigated to study the expression of TGMS-genes in the backgrounds of the three lines of hybrid rice. Pei＇ai 64S has stronger fertility restoring （Rf） genes for CMS-lines and its TGMS trait is governed by 2 pairs of independent recessive genes; The TGMS trait of 6311S is governed by a single recessive gene with weaker Rf-gene in 6311S and the TGMS trait of 360S is governed by a single recessive gene with no Rf-gene in 360S. The investigation on the fertility of F1 plants between 5 CMS-lines and 4 TGMS generations selected from F2 plants of 4 CMS-lines x 6311S confirmed that the expression of TGMS-gene was controlled by Rf-gene in the genetic background of cytoplasm of CMS-lines, but not affected by Rf-gene in the genetic background of normal fertile cytoplasm. The potential breeding strategies of TGMS-lines with cytoplasm of CMS-lines and CMS-lines with the nucleus of TGMS-genes were discussed.

Effect of Dominant Semi-Dwarf Gene on Plant Height and Its Related Traits and Sensitivity to Gibberellic Acid in Rice

Six pairs of tall and dwarf near-isogenic lines derived from a dominant semi-dwarf mutant （Y98149） were selected to study height expression and sensitivity to gibberellic acid （GA3）. The lengths of the 4-5th internode, the 3rd, 2nd, 1st internodes from the top and the panicle length in the six dwarf near isogenic lines were 97.2%, 53.3%, 65.1%, 61.9% and 94.7% of those in the six tall ones, respectively, indicating that the dominant semi-dwarfing gene significantly inhibited the internode elongation. Moreover, Y98149 （mutant type） was more sensitive to GA3 than Y98148 （wild type）, and had a lower GA3 concentration in plant, about 78% of Y98148.

Observation on Baseline Sensitivity of <i>Erysiphe necator</i>Genetic Groups to Azoxystrobin

Powdery mildew, caused by Erysiph necator, is a common and severe fungal disease of grapevine all over the world. The disease costs millions of dollars to vine growers, due to intensive use of fungicides and yield losses. Recently in population of E. necator two genetic groups have been described, the two groups seem to occupy different temporal niches, with a temporal alternation that is clear-cut in vineyards intensively treated with chemical fungicides. QoI-STAR (Quinol Outside Inhibitors-Strobilurin Type of Action and Resistance) fungicides are widely used to control the disease, and generally carry a high risk of pathogen resistance development. To clarify the behaviors of the biotrophic fungus when treated with azoxystrobin as a representative of QoI-STAR, baseline sensitivity of laboratory isolates were determined. A leaf bioassay and the primers RSCBF1 and RSCBR2 designed on the highly conserved regions of cytb gene in fungi were used. Partial sequence of E. necator cytb gene were obtained. Attempts to obtain a laboratory mutant were not totally successful. The sensitivity to azoxystrobin (EC50) in isolates of genetic group B was significantly higher than in isolates of group A, to which all the isolates collected later in the season belonged. The higher sensitivity to azoxystrobin fungicides observed in group B isolates can be at the basis of their precocious disappearance in vineyards, and can have important implications for powdery mildew control strategies.

SYNTHESIS AND pH-SENSITIVE SELF-ASSEMBLY OF DENDRITIC POLY(AMIDOAMINE)-b-POLY(L-GLUTAMATE) BIOHYBRIDS

Dendritic poly(amidoamine)-b-poly(L-glutamate)(PAMAM-b-PLG) biohybrids were synthesized by the ring-opening polymerization ofγ-benzyl-L-glutamate N-carboxyanhydride monomer,followed by the deprotection of benzyl groups on poly(benzyl-L-glutamate),and were characterized by ~1H-NMR,FT-IR and gel permeation chromatography.The self-assembly behavior of the PAMAM-b-PLG biohybrid was investigated by means of UV-Vis,dynamic light scattering (DLS),transmission electronic microscopy(TEM) and ~1H-NMR.UV-Vis analysis ...

Elongation of the Uppermost Internode for Changxuan 3S,a Thermo-Sensitive Genic Male Sterile Rice Line

Changxuan 3S, a thermo-sensitive genic male sterile （TGMS） rice line with eui gene, is derived from the TGMS rice line Pei＇ai 64S by irradiation with 350 Gy of ^60Co γ-ray. To elucidate the uppermost internode elongation of the TGMS line with eui gene, Changxuan 3S and its parent Pei＇ai 64S were used to study the effects of temperature on panicle exsertion. At 24℃, the uppermost internode of Changxuan 3S elongated the fastest from the 4^th day before flowering to 0 day （flowering）, being 2.1-fold as that of Pei＇ai 64S, whereas it elongated slowly during the 12^th day to the 4^th day before flowering and the 1^st to the 3^rd day after flowering. The uppermost internode of Changxuan 3S exserted from the flag leaf sheath at 22℃, 24℃ and 26℃, and the length of elongated uppermost internode increased with the decreasing temperatures. At 28℃, though the panicles of Changxuan 3S were still enclosed in the leaf sheath, the degree of panicle enclosure was significantly lower compared with Pei＇ai 64S. Cytological studies on Changxuan 3S showed that the uppermost internode elongation was attributed to the increase of cell number and cell elongation, and the latter was more significant. Moreover, the numbers of outermost and innermost parenchyma cells and the cell length of the uppermost internode reduced with the increasing temperatures.

Inheritance of the Male Sterility in a New Photo/Thermo-Sensitive Genie Male Sterile Line B06S of Rice

The major male sterile genes in a new photo/thermo-sensitive genie male sterile (PTGMS) line B06S of rice were analyzed by the manipulation of mixture distribution theory. The results indicated that a pair of major male sterile nuclear genes with large effects were responsible for controlling the male sterility of B06S.

Capsaicin-sensitive afferentation represents an indifferent defensive pathway from eradication in patients with H.pylori gastritis

AIM:To study the role of capsaicin-sensitive afferent nerves in Helicobacter pylori (H.pylori) positive chronic gastritis before and after eradication.METHODS:Gastric biopsy samples were obtained from corpus and antrum mucosa of 20 healthy human subjects and 18 patients with H.pylori positive chronic gastritis (n=18) before and after eradication.Tradi-tional gastric mucosal histology (and Warthin-Starry silver impregnation) and special histochemical examina-tions were carried out.Immunohistochemistry for cap-saicin receptor (TRVP1),calcitonin gene-related peptide (CGRP) and substance P (SP) were carried out by the labeled polymer immunohistological method (Lab VisionCo.,USA) using polyclonal rabbit and rat monoclonal antibodies (Abcam Ltd.,UK).RESULTS:Eradication treatment was successful in 16 patients (89%).Seven patients (7/18,39%) re-mained with moderate complaints,meanwhile 11 pa-tients (11/28,61%) had no complaints.At histological evaluation,normal gastric mucosa was detected in 4 patients after eradication treatment (4/18,22%),and moderate chronic gastritis could be seen in 14 (14/18,78%) patients.Positive immuno-staining for capsaicin receptor was seen in 35% (7/20) of controls,89% (16/18,P < 0.001) in patients before and 72% (13/18,P < 0.03) after eradication.CGRP was positive in 40% (8/20) of controls,and in 100% (18/18,P < 0.001) of patients before and in 100% (18/18,P < 0.001) after eradication.The immune-staining of gastric mucosa for substance-P was positive in 25% (5/20) of healthy con-trols,and in 5.5% (3/18,P > 0.05) of patients before and in 0% of patients (0/18,P > 0.05) after H.pylori eradication.CONCLUSION:Distibution of TRVP1 and CGRP is altered during the development of H.pylori positive chronic gastritis.The immune-staining for TRVP1,CGRP and SP rwemained unchanged before and after H.py-lori eradication treatment.The capsaicin-sensitive affer-entation is an independent from the eradication treat-ment.The 6 wk time period might not be enough time for the restituion of chronic H.pylori positive chronic gastritis.The H.pylori infection might not represent the main pathological factor in the development of chronic

鸡传染性喉气管炎病毒SYBR Green Ⅰ实时荧光定量PCR检测方法的建立与应用

鸡传染性喉气管炎病毒(Infectious laryngotracheitis virus,ILTV)以往多感染蛋鸡,近年来在蛋种鸡、商品肉鸡中连续多发,表明该病毒感染谱有扩大风险,因此有必要加强对ILTV的监测。为建立高效、灵敏的ILTV检测方法,本研究以ILTV的TK基因为靶基因设计引物,扩增并构建pMD18-T-TK质粒标准品,建立SYBR GreenⅠ实时荧光定量PCR检测方法和标准曲线,对其特异性、敏感性和重复性进行验证,并对临床疑似病例进行检测。结果发现,该检测方法特异性良好,与其他常见症状相似病原无交叉反应;最低检测浓度为1.55拷贝/μL,灵敏度是普通PCR方法的10倍;重复性好,批内、批间变异系数在0.79%~1.83%之间。表明本研究建立的ILTV实时荧光定量PCR方法特异性强、灵敏度高,可为ILTV的临床诊断和流行病学研究等提供有效的检测方法。

奥拉帕利对铂敏感复发BRCA突变卵巢癌患者PD-1/PD-L1信号通路的影响

目的探讨奥拉帕利与含铂方案对铂敏感复发乳腺癌易感基因(BRCA)突变卵巢癌(OC)患者肿瘤血管生成相关因子及程序性死亡受体-1(PD-1)/程序性死亡-配体1(PD-L1)信号通路的影响。方法回顾性选取2018年5月至2020年5月该院95例铂敏感复发BRCA突变OC患者,依据治疗方案不同分为两组,对照组47例接受紫杉类+铂类治疗,观察组48例接受奥拉帕利片。对比两组疗效、安全性、肿瘤血管生成相关因子[血管生成素-2(Ang-2)、血管内皮生长因子(VEGF)、基质细胞衍生因子-1α(SDF-1α)]、外周血CD4^(+)T细胞PD-1、PD-L1水平。结果观察组DCR比对照组高,差异有统计学意义(P<0.05)。观察组恶心、疲劳与乏力、呕吐、贫血、腹泻、白细胞减少、中性粒细胞减少及食欲减退发生率与对照组比较,差异无统计学意义(P>0.05)。观察组治疗3、6个周期血清VEGF、Ang-2及SDF-1α水平低于对照组,差异有统计学意义(P<0.05)。观察组治疗3、6个周期外周血CD4^(+)T细胞PD-1、PD-L1水平低于对照组,差异有统计学意义(P<0.05)。结论奥拉帕利应用于铂敏感复发BRCA突变OC患者中,可调控PD-1/PD-L1信号通路,降低肿瘤血管生成相关因子水平,提升治疗效果。

淮安市218批婴幼儿食品中蜡样芽胞杆菌毒力基因检测和耐药性分析

目的 了解淮安市婴幼儿食品中蜡样芽胞杆菌污染现状及其对药物敏感、主要毒力基因携带情况。方法2022~2023年抽取淮安市七个县区的婴幼儿奶粉及婴幼儿辅助食品共218批,根据GB4789.14-2014《食品安全国家标准食品微生物学检验蜡样芽胞杆菌检验》蜡样芽胞杆菌平板计数法(第一法)进行检测,并进行药物敏感性试验,采用荧光定量PCR对检出的蜡样芽胞杆菌进行溶血性(hblC)基因、非溶血性基因(nheB)、呕吐毒素基因(ces)检测。结果 218批婴幼儿食品中蜡样芽胞杆菌检出39批次,检出率达17.89%。在23种药物敏感性试验中,蜡样芽胞杆菌对庆大霉素、阿米卡星、替考拉宁、万古霉素、氯霉素、利奈唑胺、呋喃妥因、利福平、左氧氟沙星、莫西沙星、诺氟沙星敏感率100%,对环丙沙星、米诺环素、四环素、红霉素、克林霉素、莫匹罗星高浓度、复方新诺明、苯唑西林敏感率分别为97.44%、92.31%、89.74%、82.05%、38.46%、23.08%、20.51%、5.13%,对达托霉素非敏感,对头孢洛林、氨苄西林、青霉素全部耐药。溶血性hblC基因携带率高达100%,非溶血性基因nheB携带率为58.97%,本次试验未检出呕吐毒素基因ces。结论 婴幼儿食品蜡样芽孢杆菌检出率较高且检出菌中携带至少一种毒力基因的概率高达100%。对头孢洛林、氨苄西林、青霉素全部耐药,对达托霉素、苯唑西林等药物敏感性差,可为临床用药方案提供指导。

猪萨佩罗病毒聚合酶链式反应检测方法的建立及应用

猪萨佩罗病毒(Porcine sapelovirus,PSV)是引发猪腹泻的重要病毒之一。为开发一种能快速、准确检测PSV的聚合酶链式反应检测方法,针对PSV的保守区3D基因设计了一对特异性引物,优化了聚合酶链式反应退火温度,分析聚合酶链式反应检测方法特异性和灵敏度,并应用于猪场样品检测。结果表明:建立的聚合酶链式反应检测方法可特异性扩增出PSV目的片段,对其他非靶标猪腹泻病毒则无条带产生,检测低限为5拷贝/μL;在浙江地区采集的88个健康猪粪便样本中检测PSV阳性率为7.95%。该PSV聚合酶链式反应检测方法特异性强、灵敏度高,可应用于PSV前期诊断和防控。