菠菜根尖细胞有丝分裂同步化诱导

本研究以菠菜（Spinacia oleracea L.）根尖分生组织为材料，采用羟基脲（HU）和甲基胺草磷（APM）结合的双阻断法对菠菜根尖细胞进行处理，结合显微镜观察和统计学分析结果，对有丝分裂中期同步化诱导作了探讨，并对中期染色体分离也作了初步研究。在试验中分别设置了羟基脲（HU）和甲基胺草磷（APM）的不同浓度梯度，并对甲基胺草磷（APM）设置了作用时间梯度，以研究其最佳同步化诱导效果。结果表明，经1.25 mmol/L HU、10μmol/L APM处理菠菜根尖细胞，能够使细胞有丝分裂中期指数(Met.I)达40%。     

Cat2+诱导水稻细胞DNA片段化的研究

用CaCl2处理水稻愈伤组织,研究钙离子在水稻细胞凋亡过程中的作用。结果发现,水稻愈伤组织经150mmol/LcaCI2处理1~2h后,DNA无明显降解;处理3h后,DNA降解为5kb以上的大片段;处理8h后,DNA几乎完全降解成200bp~1kb的小片段。由此推测,细胞外高浓度的Ca2+对诱导凋亡细胞的DNA片段化起了重要作用。研究还发现,一定浓度的M     

以流式核型分析小麦根尖细胞的有丝分裂同步化过程

构建染色体特异文库是简化小麦基因组测序的有效途径,而通过同步化处理提高根尖细胞有丝分裂指数,使根尖细胞中富集高比例的中期染色体是实现染色体分选成功的前提。采用双阻断法用50 µmol L^-1羟基脲和100 µmol L^-1氟乐灵对普通小麦根尖细胞进行同步化处理,然后用流式核型分析法观测各周期细胞在不同时间点上的比例变化。结果显示,最佳方案为羟基脲处理10 h,恢复7 h,氟乐灵处理4 h。以此方案处理中国春小麦根尖可获得最高的有丝分裂指数,达70.1%。附加冰水处理(0°C)有助于减少染色体碎片和增加流式核型的分辨率,但对富集更多的中期染色体没有效果。     

半夏悬浮培养细胞分化形成人工种胚的同步化条件

为获得同步化的人工种胚,以半夏疏松愈伤组织为试验材料,采用悬浮培养的方法,探讨温度、咖啡因、蔗糖浓度对半夏悬浮培养细胞分化形成人工种胚的同步化作用,为半夏人工种子制作提供同步化种胚。结果表明:以MS为基本培养基,添加1.0 mg/L 2,4-D+0.5 mg/L 6-BA+30 g/L sucrose+300 mg/L CH进行悬浮培养,温度为8℃,处理12h,恢复36h,同步化效果最好,细胞分裂指数为13.86%;咖啡因浓度为2.00 mmol/L,处理2 h,恢复48 h,效果较好,细胞分裂指数为9.42%;蔗糖浓度为40 g/L,处理5 d,恢复4 d,效果一般,细胞分裂指数为5.33%。分别先后继代3次于MS+1.0 mg/L IBA+0.5 mg/L 6-BA+30 g/L sucrose+300 mg/L CH进行膨大培养及MS+0.1 mg/L IBA+0.5 mg/L 6-BA+10 g/L sucrose+300 mg/L CH进行分化培养。温度处理组微细胞团为42.33个,种胚分化率为71.11%;咖啡因处理组微细胞团为34.33个,种胚分化率为55.56%;蔗糖处理组微细胞团为40个,种胚分化率为57.78%。因此,半夏悬浮培养细胞同步化形成人工种胚的适宜条件为:MS+1.0 mg/L 2,4-D+0.5 mg/L 6-BA+30 g/L sucrose+300 mg/L CH,8℃处理12 h,恢复36 h,再转接进膨大及分化培养基继续培养。本研究为半夏人工种子生产提供技术参考。     

NaCl胁迫对留兰香基因组DNA及其甲基化的影响

盐胁迫制约农业生产的因素之一。本研究以留兰香为材料,对氯化钠处理对留兰香生长发育和生理生化反应的影响,以及基因组DNA的甲基化水平和模式进行了研究。结果表明,氯化钠处理20天后,随着盐浓度的升高,叶绿素的含量逐渐降低,MDA和脯氨酸含量逐渐增大。AFLP分析表明,处理组与对照组之间未发现特异片段,基因组序列未发生变异。MSAP分析表明,50mmol/LNaCl 处理会引起全基因组DNA的甲基化水平降低,而100和150mmol/L NaCl则诱导了全基因组DNA的甲基化水平升高;与对照相比,50、100和150 mmol/L NaCl处理后DNA甲基化和去甲基化比率分别为4.35%、9.93%、12.46%和12.73%、13.12%、20.54%。     

玉米三种不同花粉管通道法转化BcBCP1基因的初报

采用DNA柱头滴加法、花粉粒携带法和开苞叶导入法等3种不同的花粉管通道遗传转化方法,将抗逆基因BcBCP1转入3个玉米自交系合344、农大178和郑58中,对3种方法的部分影响因素进行了优化,旨在筛选出转化效率高的方法,为玉米转基因育种提供技术支持。结果表明:DNA柱头滴加法的DNA溶液浓度100g/mL,授粉后18h转化率最高。花粉粒携带法最佳处理时间为10min。开苞叶导入法在授粉后18h转化率最高,DNA浓度为100μg/mL。T0代共筛选出338株除草剂抗性植株,其中87株PCR呈阳性。DNA柱头滴加法的平均PCR阳性率最高,为1.99%;而花粉粒携带法的平均PCR阳性率最低,为0.23%;开苞叶导入法平均PCR阳性率居中,为0.32%。     

适合AFLP分析用的节瓜基因组DNA提取方法的研究

本研究以CTAB法为基础,采用以下5种处理提取节瓜叶片基因组DNA:A、PVPP(3%)+抗坏血酸(100 mmol/L);B、β-巯基乙醇(3%)+抗坏血酸(100 mmol/L):C、PVPP(3%)+半胱氨酸(5%);D、PVPP(3%)+β-巯基乙醇(3%);E、抗坏血酸(100 mmol/L)+半胱氨酸(5%).实验结果表明:在5种处理中,C处理提取的DNA质量最高,其溶液呈无色透明状,经核酸蛋白仪测定其OD260/OD280为1.8～2.0;经0.8%琼脂糖凝胶电泳检测,节瓜叶片基因组DNA带型清晰、完整、无降解,蛋白、多糖、酚类及RNA等去除比较彻底;经AFLP分析,酶切彻底,PCR扩增后的带型清晰稳定,重复性好,说明C处理提取的节瓜基因组DNA完全满足AFLP分析的要求.     

外源钙离子对高温胁迫下草莓幼苗影响的转录组分析

为探究添加外源钙离子后高温胁迫下草莓幼苗叶片的基因表达变化情况,本试验以‘红颜’草莓幼苗为材料,设置4个试验处理,分别为对照(昼/夜25℃/17℃16 h/8 h)每株喷施20 mL蒸馏水、对照每株喷施20mL的8mmol/L CaCl₂溶液、高温(昼/夜42℃/34℃16h/8h)每株喷施20mL蒸馏水、高温每株喷施20 mL的8 mmol/L CaCl₂溶液,在高温胁迫第3天时取叶片进行转录组测序和生物信息学分析。结果表明：与高温喷施蒸馏水处理组相比,高温喷施外源钙离子处理组发生差异表达的基因共有2 452个,其中1 504个上调表达,948个下调表达;GO富集分析显示在细胞组成中,差异基因显著富集在细胞、细胞部分和细胞器;在分子功能中显著富集的条目是结合、催化活性、结构分子活性;显著富集的生物过程为代谢过程、细胞过程、刺激应答等;KEGG富集分析表明显著富集的代谢通路有植物与病原体的相互作用和MAPK植物信号传导途径-植物和植物激素信号转导等,进一步分析发现添加外源钙离子会使一些编码钙离子感受蛋白、WRKY转录因子、MYB转录因子、热激...     

VHZ对玉米分生组织毒害的细胞及生化分析

研究了微管解聚型三唑类化合物VHZ对玉米分生组织的影响,从细胞学及生物化学水平分析了VHZ的毒害机理.结果表明,VHZ能够诱导玉米根尖分生组织细胞前期、中期的同步化以及染色体变异(染色体桥、染色体滞后、染色体凝集).临界浓度和时间分别为20 μmol/L和12 h.免疫荧光标记检测表明,同步化细胞的形成以及染色体结构变异与纺锤丝的功能受到抑制有关.2D/SDS-PAGE检测发现,经VHZ处理后玉米根尖蛋白有两种组分消失,四种蛋白质组分含量有不同程度的降低,这些蛋白质组分可能对VHZ的防御机制有重要作用.     

硫化氢对白桦悬浮细胞生长、黄酮和多酚累积的影响

旨在探究硫化氢(H2S)对白桦悬浮细胞生长、黄酮和多酚等次生代谢物累积的影响。将H2S供体硫氢化钠(NaHS)添加到培养8 天的白桦悬浮培养体系中,采用pH计、电导率仪和紫外分光光度计等分析pH值、电导率、细胞活力、丙二醛、过氧化氢酶(CAT)和苯丙氨酸解氨酶(PAL)活性、黄酮和多酚含量。NaHS处理后白桦悬浮细胞培养液的pH值呈降低趋势、电导率呈增加趋势;白桦悬浮细胞中的丙二醛含量、CAT和PAL酶活性、黄酮和多酚含量呈增加趋势,而细胞活力基本呈下降趋势,且NaHS处理对上述白桦悬浮细胞生长、黄酮和多酚含量的影响存在浓度和时间效应。其中,1 mmol/L NaHS处理24 h 时黄酮和多酚含量最高,分别是对照组的128.57%和136.89%。NaHS处理促进了白桦悬浮细胞中黄酮和多酚的累积。     

腐胺对白桦悬浮细胞中激素和黄酮积累影响的初步研究

为明确腐胺对白桦悬浮细胞中激素和黄酮累积的影响,在白桦悬浮细胞生长的第8 天添加不同浓度的腐胺、脱落酸(ABA)、茉莉酸甲酯(MeJA)、生长素(IAA)和玉米素核苷(ZR),采用紫外分光光度计法分析白桦悬浮细胞中黄酮含量和利用酶联免疫方法测定ABA、JA、IAA和ZR含量。外源腐胺和激素的添加均不同程度地提高了白桦悬浮细胞中黄酮的累积量,其中1 mmol/L 腐胺处理24 h 时增幅最高,增加了97.90%;激素中1 mg/LABA处理后增幅最高,增加了103.40%。腐胺和激素共同处理后黄酮含量的增幅均较其单独处理提高。其中1 mg/L 腐胺与5 mg/L JA 处理后黄酮含量增幅最高,比对照和腐胺处理分别增加了197.43%和103.57%。同时腐胺处理后增加了IAA 和ZR的含量,而降低了ABA和JA 含量。进一步分析其相关性发现,腐胺处理后激素、激素比与黄酮含量的相关性增幅不同,其中JA与黄酮含量的相关性增幅最大,由对照的-0.883 变为0.903。腐胺与ABA、JA、IAA和ZR协同促进了白桦悬浮细胞中黄酮的累积。     

ISSR Analysis of Genetic Background of Triploid Female and Diploid Male of Siraitia grosvenorii

[Objective] To evaluate the genetic background of triploid female and diploid male of Siraitia grosvenorii and to provide biological references for fine varieties breeding of seedless S. grosvenorii. [Methods]Inter simple sequence repeats(ISSR) marker was developed to analyze the genetic background among 28 samples of S. grosvenorii,and cluster analysis and double principal coordinate analysis were revealed by the NTSYS-pc software and Gen AIEx software,respectively. [Results]13 ISSR primers selected out of 100 primers for amplification,and a total of 131 unambiguous bands were obtained,among which 99(PPB = 75. 57%) were polymorphic. The results of cluster analysis and double principal coordinate analysis showed that there was a certain richness of genetic background in triploid female and diploid male of S. grosvenorii. But the genetic similarity coefficients of majority were relatively great with close genetic distance. [Conclusions] The triploid female and diploid male of S. grosvenorii had relatively low complexity of genetic background,so that germplasm innovation strategies should be carried out to enrich the genetic background of seedless S. grosvenorii.     

山丹丹原生质体的分离条件探究

[目的]本研究旨在建立山丹丹高效原生质体分离体系,为山丹丹种质资源的有效利用提供科学依据。[方法]以山丹丹鳞茎胚性愈伤组织为材料,通过酶解法获得山丹丹原生质体,进一步采用3因素4水平L16(3⁴)正交试验 。[结果]结果表明：(1)在预处理条件下(0.015 g/mL纤维素酶+0.005 g/mL果胶酶+0.65 mol/L甘露醇)得出最佳酶解时间为6 h,产量为3.47×10⁶个/ mL；(2)对正交试验结果进行极差分析,确定原生质体产量主次因素为：纤维素酶>果胶酶浓度>甘露醇浓度,同时得到最佳酶组合为0.02 g/mL的纤维素酶浓度、0.008g/mL的果胶酶浓度和最佳渗透压浓度为0.6 mol/L甘露醇,原生质体产量最高为3.47×10⁶个/ mL,活力为60.32%；(3)在最佳酶组合处理下,采用暗处理可将原生质体存活率提高到76.15 %；在15 min、1000 r/min时,原生质体产量最高,为3.51×10⁶/mL；在10 min、800 r/min时,原生质体活力最高,为61.2 %。 [结论] 该试验获得了山丹丹原生质体最佳分离条件,建立原生质体分离体系,为山丹丹种质资源的有效利用提供研究奠定基础。     

Genetic diversity in soybean genotypes from north‐eastern China and identification of candidate markers associated with maturity rating

Random amplified polymorphic DNA (RAPD) and simple sequence repeat (SSR) markers were used to estimate the genetic relationships among 101 soybean cultivars developed in north‐eastern China. Fifty‐three fragments of the 100 RAPD markers and 35 SSR markers tested were polymorphic across the 101 soybean cultivars. Similarity values among these soybean cultivars ranged from 45.2% to 100% for RAPD data, and ranged from 36.1% to 100% for SSR data. The similarity matrices for SSR data and RAPD data were moderately correlated (r = 0.31, P < 0.05). Cluster analyses indicated that the cultivars released from the same seed company were mostly grouped together. A principal component analysis, based on the combined RAPD and SSR data, yielded a good separation of soybean varieties with different maturity ratings [represented by soybean Heat Unit (HU)]. The varieties with HU < 2200 were well separated from those with HU > 2200. Four RAPD markers and eight SSR markers were significantly associated with the maturity ratings of soybean.     

南美蟛蜞菊毛状根乙醇提取液对种子萌发的影响

摘 要：【研究目的】本文拟通过用不同浓度的蟛蜞菊毛状根的乙醇提取液浸泡三叶鬼针草(Bidens pilosa L.)等种子研究其化感作用,为今后开展通过南美蟛蜞菊离体培养毛状根来生产高效低毒的“生物农药”及其应用奠定实验和技术基础。【方法】我们用0.5 g/mL、1.0 g/mL两种浓度蟛蜞菊毛状根的乙醇提取液,分别浸泡三叶鬼针草(Bidens pilosa L.)等种子3h、12h和24 h后,进行种子萌发实验。以清水浸泡后的种子萌发情况作为对照相比。【结果】提取液的浓度越高或浸泡时间越长,种子的萌发率越低,而且发现在0.5 g/mL(鲜重)的低浓度提取液中浸泡12 h以上,鬼针草、马唐和狗尾草已经不能正常萌发,在1.0 g/mL(鲜重)的高浓度提取液中浸泡12 h以上对种子的抑制作用更明显,鬼针草、马唐、稗草、狗尾草、蒲公英种子均不能萌发。而且发现无论是用0.5 g/mL(鲜重)的低浓度提取液还是用1.0 g/mL(鲜重)的高浓度提取液进行浸泡,浸泡时间在24 h以上,抑制效果就更加明显。【结论】结论：蟛蜞菊毛状根具有较强的化感作用。     

Synchronization and Detection of Rat Hepatocellular Carcinoma Cell Line Cells

The proliferation and division of cells are the most basic reaction. The effects of this reaction are the fact that the cell cycle was been influenced. Normal cells will change into tumor cells when its cell cycle running out of control; on the other hand, it will become feeble and die when its cell cycle stopping. Methods of synchronization and detection with flow cytometer of rat hepatocellular carcinoma cell line (HTC) are established in our laboratory. The results show that: as HTC cell is in the logarithmic period, the synchronous 72.10% G 1 phase and 98.94% S phase cells are achieved through thymine 2 desoryriboside and cochicine sequential blockage method and double thymine 2 desoryriboside blockage method respectively. It is proved that those results ware stable and reliable.     

葡萄籽原花青素对大鼠脾脏淋巴细胞与肝细胞增值的影响

为了研究葡萄籽原花青素(Procyanidin from Grape Seed,GSP)对大鼠脾脏淋巴细胞与肝细胞增值与活化的影响,揭示GSP提高机体免疫性能的作用机理,为后续的试验分组提供理论依据。试验采用不同终浓度的GSP培养液[0.00 （对照）、0.05、0.10、0.50、1.00、5.00、10.00、15.00]与大鼠脾脏淋巴细胞, [0.00 （对照）、0.05、0.10、0.50、1.00、5.00、10.00、15.00、20.00、25.00 μg/mL ]与大鼠肝细胞系IAR-20共育。在12、24、48 h后,用酶标仪在490 nm处测得各孔的OD值。结果显示：（1）添加0.05~5.00 μg/mL的GSP促进淋巴细胞增值,当浓度超过10.00 μg/mL将抑制淋巴细胞增值。（2）添加0.05~10.00 μg/mL 的GSP促进肝细胞增值,当浓度超过20.00 μg/mL 将抑制肝细胞增值。（3）一定范围内,GSP对淋巴细胞和肝细胞的促增值作用有浓度和时间依赖性。葡萄籽原花青素能够促进体外培养大鼠的脾脏淋巴细胞和肝细胞增值活性,具有提高细胞抗氧化和细胞免疫的作用。     

Effects of Medicated Serum of HERBA POLYGONI PERFOLIATI on HBsAg and HBeAg Secretion in 2215 Cells

[Objective] To research the effects of medicated serum of HERBA POLYGONI PERFOLIATI on HBsAg and HBeAg secretion in2215 cells. [Methods]Destruction rate of medicated serum of HERBA POLYGONI PERFOLIATI to 2215 cells and HL-7702 were detected by MTT method; HBsAg and HBeAg secretion in cell supernatant was detected by ELISA method when four different concentrations of medicated serums were applied in 2215 cells for 72 h. [Results]20% and 40% medicated serums showed inhibitory effects on HBsAg and HBeAg secretion,showing certain dose-effect relationship. At the same time,medicated serum of HERBA POLYGONI PERFOLIATI had less significant inhibitory effects on HL-7702 than that on 2215 cells. [Conclusions] Medicated serum of HERBA POLYGONI PERFOLIATI had certain toxic and anti-HBV effects in vitro.     

In Vitro Bacteriostasis of Extracts from Gingko biloba L.

[Objective]To discuss the in vitro bacteriostasis of the extracts from the seeds and leaves of Gingko biloba L. [Methods]Crude extracting solutions were extracted from the G. biloba leaves and seeds by ethanol extracting method. Inhibitory effects of the extracting solutions of G. biloba leaves and seeds on Escherichia coli,Proteus vulgaris,Bacillus subtilus and Staphylococcus aireus were researched. [Results]Extracts from G. biloba leaves had relatively strong effects on S. aireus with the minimal inhibitory concentration being 125. 00 mg/mL,and had certain inhibitory effects on E. coli with the minimal inhibitory concentration being 250. 00 mg /mL. However,it had no inhibitory effects on B. subtilus or P. vulgaris. Extracts from G. biloba seeds had no inhibitory effects on four bacteria. [Conclusions] Extracts from G. biloba leaves had relatively good inhibitory effects on E. coli and S. aireus,but had no effects on B. subtilus or P. vulgaris. Extracts from G. biloba seeds had no inhibitory effects on all the four bacteria.