间接原位RT-PCR法检测辣椒叶组织中辣椒轻斑驳病毒的分布

为了了解辣椒轻斑驳病毒（Pepper mild mottle virus,PMMoV）在辣椒组织中的分布特点,探索利用间接原位RT-PCR技术对病毒在辣椒组织中进行定位。利用改进的CTAB法从辣椒叶片中提取总RNA,利用RT-PCR技术扩增出PMMoV的特异片段,并经克隆测序,证明其为PMMoV的特异片段。利用PCR技术,制备出了地高辛标记的特异cDNA探针。对原位RT-PCR反应体系进行优化,经过切片制备（切片厚度7 μm、应用Superfrost plus正电荷防脱载玻片）、预处理（1 mg ? L-1蛋白酶K消化5 min）、RT-PCR（37 ℃反转录2.5 h、PCR退火温度为58 ℃）、杂交检测,建立了应用间接原位RT-PCR技术检测PMMoV在辣椒组织中分布的方法。原位RT-PCR结果显示,叶片组织中PMMoV阳性信号主要分布于栅栏组织,其次是海绵组织,表皮组织中也有少量阳性信号。此外,叶柄中可见少量阳性信号,主要位于表皮组织。     

树莓品种‘Kivigold’快繁技术体系建立

以树莓（Rubus idaeus）品种‘Kivigold’带腋芽茎段为外植体,研究消毒剂HgCl2（0.1%）不同的消毒时间对外植体培养的影响,对不定芽增殖培养基和生根培养基中适宜的激素种类及浓度、生根苗移栽驯化中适宜的基质进行了筛选。结果显示,树莓品种‘Kivigold’外植体的最佳消毒时间为10 min,初代培养时只需添加6-BA即可满足外植体萌发和生长的需要;在不定芽增殖过程中,细胞分裂素主要影响不定芽的增殖,而生长素主要影响不定芽的生长,以质量浓度低于1.5 mg/L的6-BA以及质量浓度低于0.5 mg/L的NAA较为适宜,3种碳源中蔗糖更有利于不定芽的增殖和生长;经过进一步的筛选,确定适宜于‘Kivigold’不定芽增殖和生长的培养基为MS+1.00 mg/L 6-BA +0.10 mg/L NAA（含20 g/L蔗糖和5.9 g/L琼脂,pH 5.8）,适宜于‘Kivigold’不定芽生根的培养基为1/2 MS+0.10 mg/L NAA（含20 g/L蔗糖和5.9 g/L琼脂,pH 5.8）;在移栽驯化中最适宜的栽培基质为泥炭土:珍珠岩=1:1,移栽成活率可达到93.33%。     

高加索三叶草×白三叶胚萌发条件的探索

以六倍体高加索三叶草Trifolium ambiguum与四倍体白三叶T.repens为亲本,通过远缘杂交,采用组织培养手段对杂交胚的萌发条件进行探索.结果表明:以高加索三叶草为母本、白三叶为父本,在花期8:00-9:00授粉,授粉后12～13 d取胚,用质量分数10%NaClO灭菌6.5～7 min,在不加任何外源激素的改良MS培养基上,胚萌发率最高,达8%.高频扩繁时,再附加0.25 mg/L 2,4-D+0.50 mg/L 6-BA可获得较高萌发率.     

白三叶基因工程改良研究进展

白三叶（Trifolium repens L.）作为优良牧草和地被植物具有重要的应用价值。但其耐盐碱、干旱能力差等原因致使应用受到广泛限制,转基因技术的快速发展为白三叶种质创新提供了有效技术手段,为开展白三叶分子育种奠定了基础。本文就近30年来有关白三叶组织培养、转化方法、转基因遗传改良方面的研究进展进行了综述,并对其应用进行了展望。     

In vitro expression and inhibition of procoagulant activity produced by bovine alveolar macrophages and peripheral blood cells

Local and systemic activation of coagulation is frequently associated with bacterial sepsis. The coagulopathy is due, at least in part, to expression of tissue factor (TF) by monocytes and macrophages. The purpose of this study was to evaluate the expression of procoagulant activity by bovine alveolar macrophages, leukocytes and platelets, and to determine the relative potency of three chemical inhibitors of TF expression (pentoxifylline, retinoic acid, and cyclosporin A). Bovine alveolar macrophages were stimulated with lipopolysaccharide (LPS) derived from Pasteurella haemolytica or recombinant bovine tumour nervous factor (TNF) and dose- and time-dependent effects on TF expression were studied. LPS and TNF induced TF expression in alveolar macrophages and LPS treatment of whole blood induced TF expression in mononuclear cells. Neutrophils and platelets also expressed procoagulant activity, but this activity was not inhibited by anti-bovine TF monoclonal antibody. Pentoxifylline (40 mol/L), retinoic acid (0.01 mmol/L) and cyclosporin A (0.08 mol/L) inhibited TF expression when added concurrently with LPS or TNF, but not when added 4 h after stimulation. TF mRNA was not detected in unstimulated alveolar macrophages by Northern blot analysis. In contrast, exposure to LPS or TNF for 6 h induced marked expression of TF mRNA, which was inhibited by treatment with pentoxifylline, retinoic acid and cyclosporin A. Expression of TNF by alveolar macrophages stimulated with LPS was also inhibited by these compounds. Our results indicate that procoagulant activity expressed by alveolar macrophages and monocytes is associated with expression of TF, whereas procoagulant activity expressed by neutrophils and platelets is not. The concentrations of pentoxifylline and retinoic acid necessary for inhibition of TF expression in vitro may not be achievable in vivo owing to their toxic effects. However, the in vitro concentration of cyclosporin A that inhibited TF expression did not exceed the plasma concentration observed in humans, and therefore may be useful for inhibition of TF expression in vivo.Abbreviations BAL bronchoalveolar lavage - LPS lipopolysaccharide - cDNA cloned deoxyribonucleic acid - cAMP cyclic adenosine monophosphate - GAPDH glyceraldehyde phosphate dehydrogenase - mRNA messenger ribonucleic acid - TF tissue factor - TNF tumour necrosis factor - DPBS Dulbecco's phosphate-buffered saline     

独蒜兰属植物研究现状

从地理分布、形态特征、生态学、繁殖栽培和资源开发等方面,对独蒜兰的研究进展进行了综述,为独蒜兰的进一步研究和开发利用提供了参考。     

狭叶三七茎段芽试管苗培养研究

为保护野生资源,以狭叶三七具有生长点的茎段为材料,进行生长芽培养、生长芽试管苗培养、试管苗茎段繁殖培养,以及试管苗移栽、定植的研究。结果证明：1／2MS＋蔗糖20g／L＋6-BA0．2mg／L＋NAA0．1mg／L＋IAA0．1mg／L是生长茅培养的适宜培养基。1／2MS＋蔗糖16g／L＋IAA0．2mg／L＋NAA0．1mg／L是试管苗培养和试管苗继代繁殖培养的适宜培养基。试管苗移栽成活率为98．1％,定植成活率为98．9％,定植成活的试管苗保持了野生狭叶三七植物学性状。     

Evidence for functional G-coupled protein receptors 43 and 120 in subcutaneous and intramuscular adipose tissue of Angus crossbred steers

We conducted 3 independent experiments to demonstrate functional G-coupled protein receptor 43 (GPR43) and GPR120 in bovine intramuscular (i.m.) and subcutaneous (s.c.) adipose tissues. We hypothesized that media volatile fatty acids and long-chain fatty acids would affect cAMP-activated protein kinase-alpha (AMPKα) protein expression and cAMP concentrations differently in i.m. and s.c. adipose tissue. Experiment 1: oleic acid (18:1n-9) decreased phosphorylated AMPKα protein (p-AMPKα) and the p-AMPKα/AMPKα protein ratio in i.m. preadipocytes, increased the p-AMPKα/AMPKα protein ratio in bovine satellite cells, and had no effect in s.c. preadipocytes. Experment 2: ex vivo explants from the 5th to 8th longissimus thoracic rib muscle section of Angus crossbred steers were cultured 48 hr in media containing 0.25 µM ciglitizone, 5 mM glucose, and 5 mM acetate, in the absence or the presence of 100 µM oleic acid. Oleic acid increased acetate incorporation into fatty acids and GPR43 gene expression in i.m. adipose tissue (P < 0.05), but oleic acid had no effect on fatty acid synthesis or GPR43 expression in s.c. adipose tissue. Experiment 3: fresh s.c. and i.m. adipose tissue from the 5th to 8th longissimus thoracic rib muscle section of Angus crossbred steers was transferred immediately to 6-well culture plates containing 3 mL of KHB/Hepes/5 mM glucose. Samples were preincubated with 0.5 mM theophylline plus 10 μM forskolin for 30 min, after which increasing concentrations of acetate or propionate (0, 10⁻³, 10^−2.3, and 10⁻³ M) in the absence or the presence of 100 μM oleic acid or 100 µM palmitic acid (16:0) were added to the incubation media. Acetate had no effect on forskolin-stimulated cAMP production in s.c. adipose tissue but decreased cAMP in i.m. adipose tissue (P < 0.05); this indicates a functional GPR43 receptor in i.m. adipose tissue. The combination of 10⁻² M acetate and oleic acid decrease cAMP production in s.c. adipose tissue, consistent with GPR120 receptor activity, but oleic acid and palmitic acid attenuated the depression of cAMP production caused by acetate in i.m. adipose tissue. Palmitic acid depressed cAMP production in s.c. adipose tissue, and increased cAMP production in i.m. adipose tissue (P < 0.05). Propionate had no effect on cAMP production in s.c. or i.m. adipose tissue. These results provide evidence for functional GPR43 receptors in i.m. adipose tissue and GPR120 receptors in s.c. adipose tissue, both of which would suppress lipolysis.     

TDZ对杏香兔耳风组培苗复壮的影响

为了了解TDZ对组培苗增殖的影响情况,探讨了TDZ在衰退组培苗复壮过程中的作用,以增殖衰退的杏香兔耳风组培苗为试验材料,就TDZ的不同添加方式(抽滤灭菌和高压灭菌)和不同浓度(0.0、0.2、0.4、0.6、0.8、1.0 mg/L)处理对组培苗增殖系数、有效芽及苗高生长的影响情况进行了试验。结果表明:当TDZ的浓度0.2 mg/L时,TDZ的添加方式对组培苗的增殖系数和有效芽数均有较大影响,抽滤灭菌后的增殖系数和有效芽数要明显高于高压灭菌的;不同浓度的TDZ对有效芽数和髙生长的影响均有极显著的差异性(P0.01),当TDZ浓度为0.8 mg/L时,苗质量总体达到最优,增殖系数为5.11,达到了预期的复壮效果;最优生根培养基配方为1/2MS+NAA 0.02 mg/L+IBA 0.1 mg/L。在杏香兔耳风组培苗复壮过程中,TDZ可作为一种有效的植物生长调节剂使用。     

牡丹组织培养中褐化的发生原因与防止方法的研究

在牡丹组织培养过程中,褐化现象普遍发生,并影响着牡丹培养物的正常生长与增殖。因此通过改变培养条件、培养方式以及在培养基中添加抑褐剂的方法,对品种为青龙卧墨池的牡丹组培物褐化的防止进行了研究,结果表明:添加生长素会加剧启动培养中外植体的褐化程度,暗培养对愈伤组织抑制褐化很有效,悬浮培养未能减缓褐化却能使愈伤快速增殖。吸附剂聚乙烯吡咯烷酮(PVP)和结冷胶(Phytagel)能够有效地缓解褐化。抑褐剂硫代硫酸钠(Na2S2O3)、抗坏血酸(VC)和铜试剂(DDTC)在牡丹的组培中加剧了组织培养物的褐化程度。