诱导

0Mg/L 6-benzylaminopurine was the optimal concentration of shoots quantity inducing which frequency of inducing shoots and number of shoots per explant were 100% and 11.1 respectively. 0.5mg/L 6-benzylaminopurine was the optimal concentration of shoots length inducing which average length of shoots was 12.0mm. Explant size had significantly effects on direct shoots quantity and length induction. When complete cotyledon used as explant, frequency of inducing shoots, number of shoots per explant and average length of shoots could approach to 100%, 6.7 and 17.1mm respectively. 2.0mg/L silver nitrate could significantly increase shoot quantity induction, both frequency of inducing shoots and number of shoots per explant could be as high as 97.2% and 4.2 respectively. But silver nitrate also could restrain shoots elongation.

结果表明：不同黄瓜品种直接不定芽数量诱导存在显著差异，但长度诱导无显著差异；6-苄氨基嘌呤对直接不定芽数量和长度的诱导作用显著，其中4.0mg/L为数量诱导适宜浓度，其出芽率和每外植体出芽数达到最高，分别为100.0%和11.1，0.5mg/L为长度诱导适宜浓度，其平均芽长12.0mm；外植体的大小对直接不定芽数量和长度均具显著影响，随着子叶的增大诱导直接不定芽的数量和长度增加，其中单片完整子叶为最适宜大小，其出芽率、每外植体出芽数和平均芽长分别达到100%、6.7和17.1mm；硝酸银也对直接不定芽数量诱导影响显著，其中2.0mg/L为适宜浓度，出芽率和每外植体出芽数分别达到97.2%和4.2，但对芽长有抑制作用。

B Of the six basic media MS, MS1/2 (half-strength of MS salts and vitamins), WPM, DKW, B5 and SH, MS1/2 was the most proper one to induce somatic embryos. Somatic embryos generally regenerated directly from excised zygotic cotyledons. PGRs combination affected somatic embryogenesis significantly. Medium with NAA 1mg/L, TDZ 0.05mg/L, IBA 2—10mg/L combined with BA 10mg/L, or IBA 10mg/L integrated with BA 0-2mg/L gave the highest induction rate. Excised zygotic hypocotyls had the strongest potential to produce callus. Callus induction was also affected significantly by media and PGRs. The proper callus induction condition was MS1/2 medium containing NAA 1mg/L, IBA 10mg/L, BA 2-5mg/L and TDZ 0.05mg/L. Harvest period affect somatic embryogenesis significantly. Zygotic embryo explants collected from the end of July to the middle of August had strong potential to generate somatic embryos, when endosperm finished solidification, different parts of the embryos were completely formed, the size of embryos occupied about 2/3 of the embryo sac. Provided with optimized conditions, direct somatic embryogenesis rate can attain to 33. 68%, and callus induction rate of hypocotyls was up to 90.7%. Cytological observation on megasporogenesis and zygotic embryogenesis of Manchurian ash showed that the ovary was twicarpellum, twilocular with two ovules each loculus. The ovule was tenuinucellar and anatropous, with one megasporcocyte. The development of embryo sac is of the Polygoum type.

体细胞胚胎发生研究的结果表明：(1)成熟过程中的合子胚是诱导水曲柳体细胞胚胎发生的最佳外植体材料；(2)在所试验到的MS、MS1/2（将MS的所有成分均减半）、WPM、DKW、B〓、SH等六种基本培养基中，MS1/2是最适合诱导水曲柳体细胞胚胎发生的基本培养基；(3)水曲柳的体细胞胚胎发生以直接发生为主，体细胞胚主要来自于从合子胚分离的完整子叶；(4)培养基中的激素组合对水曲柳的体细胞胚胎发生有显著影响，诱导直接体细胞胚发生较好的激素组合有NAA 1mg/L+IBA 2，5，10mg/L+BA 10mg/L+TDZ 0.05mg/L和NAA 1mg/L+IBA 10mg/L+BA 0，2mg/L+TDZ 0.05mg/L；(5)合子胚分离的下胚轴具有最强的愈伤组织诱导潜力，少数愈伤组织可以分化出体细胞胚；(6)愈伤组织的诱导也受培养基和激素配比的显著影响，最适宜诱导的培养条件为MS1/2+NAA 1mg/L+IBA 10mg/L+BA 2，5mg/L+TDZ0.05mg/L；(7)采种时间对体细胞胚胎发生有显著影响。7月末到8月中旬的合子胚具有较强的体细胞胚发生潜力，此时种子尚未成熟，胚乳已呈固态，种胚的各个部分已分化完全，种胚体积占胚腔的大约2/3；(8)在各自综合的最适条件下，完整子叶的体细胞胚诱导率可达33.68%，下胚轴的愈伤组织诱导率可达90.7%。

The major results of this proposal are as follows:(1) ABA induces a rapid, substantial accumulation of apoplastic H2O2 in mesophyll and bundle sheath cells of maize leaves, and the accumulation of apoplastic H2O2 is involved in the induction of the chloroplastic and cytosolic antioxidant enzymes.(2) ABA-induced H2O2 production activates a 46 kDa MAPK, which in turn induces the expression of antioxidant genes and up-regulates the activities of antioxidant enzymes. The activation of MAPK also enhances H2O2 production, forming a positive amplification loop.(3) Water stress also induces the activation of a 46 kDa MAPK, which is dependent on the accumulation of ABA and H2O2 production induced by water stress and involved in the up-regulation of the expression and the activities of antioxidant enzymes.(4) ABA-induced H2O2 production mediates NO generation, which in turn activates a 46 kDa MAPK and results in the up-regulation in the expression and the activities of antioxidant enzymes in ABA signaling. NO-independent signaling is also involved in ABA- and H2O2-induced antioxidant defense.

本项目的主要研究结果如下：（1）ABA诱导的H2O2产生主要出现在叶肉细胞与维管束鞘细胞的质外体中，质外体H2O2的积累能够上调叶绿体与细胞溶质中抗氧化酶的活性；（2）ABA诱导的H2O2产生活化一个46kDa的MAPK，由此而导致编码抗氧化防护酶基因的表达与酶活性的上调；而MAPK的活化也能增强H2O2的产生，从而形成一个正的反馈调节环；（3）水分胁迫也能诱导一46kDa MAPK活化，这一MAPK活化依赖于水分胁迫诱导的ABA积累以及H2O2的产生，同时参与水分胁迫诱导的抗氧化防护基因的表达与抗氧化酶活性的上调；（4）ABA诱导一个依赖于H2O2的NO产生，NO活化一个46kDa的MAPK，从而导致抗氧化防护基因的表达以及抗氧化酶活性的上调；同时一个不依赖于NO的信号转导途径也存在于ABA诱导的抗氧化防护过程中。

Results The peak of antibacterial activity appeared at 36-48 hafter induced by four inducements (Because many M.domestica larvae pupated,we did not observe the peak of antibacterial activity by heat-shock). The antibacterial activity of antibacterial proteins by S.aureus was best,and the antibacterial activity of antibacterial proteins by heat-shock was better than control group.

结果 各诱导组诱导家蝇幼虫产生抗菌蛋白达最大活性时间为诱导后36～48h(热诱导后家蝇幼虫很快化蛹，未观察抗菌蛋白产生高峰时间)；金黄色葡萄球菌诱导后的抗菌蛋白抗菌活性最佳，热诱导后的抗菌蛋白抗菌活性也较未诱导组好。

Results are as followed:1 Exposure of HELF cells to BP caused c-Jun activation,and increased the activity of MAPK,PI-3K,p53 and cyclin D1 pathway.2 BP-induced c-Jun activation was inhibited by dominant negative mutants of extracellular signal-regulated protein kinase or c-Jun NH_2-terminal kinase,but not by p38,impling that JNK and ERK pathways medicate c-Jun activation induced by BP.3 Overexpression of dominant-negative mutants PI-3K and Akt potently blocked phosphorylations of c-Jun and ERK,but not JNK in response to BP,suggesting that PI-3K/Akt pathway positively regulates BP-induced c-Jun activation through ERK.4 Inhibition of p53 by its chemical or molecular inhibitor markedly increased the phosphorylation levels of c-Jun,Akt and ERK upon BP stimulation,indicating that p53 negatively medicates BP-induced c-Jun activation through PI-3K/Akt/ERK pathway.5 The cell lines expressed TAM67 exhibits no significant affecting normal cell growth properties.6 TAM67 was able to significantly block G_1-S transition and subsequent cell proliferation,suggesting that c-Jun is essential for cell cycle alternations elicited by BP.7 Overexpression of TAM67 impaired BP-induced cyclin D1 activation,decreasing expression of E2F1 and pRb,indicating that c-Jun participates in the modulation of BP-induced activation of cyclin D1/pRb/E2F1 pathway.8 Stably expression of TAM67 led to the increases in the expression levels of p53 and p21,elevating phosphorylation level of p53,clearly indicating that c-Jun regulates p53/p21 pathway activation induced by BRCollectively,PI3K/Akt/ERK pathway mediated BP-induced c-Jun activation through p53-dependent mechanism.

结果显示：1BP刺激细胞可促进c-Jun活化，并伴随着MAPK、PI-3K、p53和cyclinD1通路各组成成分的活性增强。2利用MAPK通路的显性失活突变体分别阻断细胞外信号调节激酶和c-Jun氨基末端激酶活性，均可明显抑制BP诱导的c-Jun活化，但阻断p38活性对BP引起的c-Jun活化无明显影响，提示JNK和ERK通路参与调控BP诱导的c-Jun活化。3过表达PI-3K和Akt的显性失活突变体也可显著抑制BP诱导的c-Jun活化，并降低磷酸化ERK的表达水平，但对磷酸化JNK的表达水平无明显影响，说明PI-3K/Akt通路通过ERK正性调控了BP诱导的c-Jun活化。4p53的化学/分子抑制剂能使BP作用的细胞内c-Jun活性明显增加，并同时诱导Akt和ERK的磷酸化水平的升高，表明p53可通过PI-3K/Akt/ERK通路对BP诱导的c-Jun活化进行负性调控。5随后观察转染细胞的生长情况，发现TAM67对细胞正常生长和形态无明显影响。6稳定表达TAM67可有效抑制BP诱导的S期细胞数的增加，提示c-Jun在BP致细胞周期改变的过程中发挥了重要作用。7TAM67过表达能够抑制BP诱导的cyclin D1活化，降低磷酸化Rb以及E2F1蛋白表达水平，表明c-Jun参与调控BP诱导的cyclin D1/Rb/E2F1通路的活化。8过表达TAM67可使BP刺激的细胞中p53、p21总蛋白以及p53磷酸化的表达水平明显升高，可见c-Jun也参与调控BP诱导的p53/p21通路活化。

New reactions found in this program include: photoinduced regioselective free radical addition to aromatic imines;photoinduced Dielse-Alder reactions of aromatic imines for synthesis of tetrahydroquinoline derivatives;Photoinduced radical cation mediated selective ring opening reaction of epoxides;Formation of novel macrocyclic crown ethers by photoinduced reaction of diazonaphthaquinones;photoinduced selective desilylation of silyl ethers.

用这种方法成功的发现了下列在有机合成中有意义的新反应：(1)光诱导产生的自由基正离子诱导的自由基对芳香亚胺的区域选择性加成反应；(2)用光诱导的芳香亚胺的Diels-Alder反应合成四氢喹啉衍生物；(3)经光诱导产生的自由基正离子基诱导的环氧化物选择性开环反应；（4）重氮萘酮在环醚中经光诱导反应形成新奇的大环冠醚；（5）硅醚的光诱导选择性脱硅基反应。

ANP and CX43 began to express at 2nd week after induction and increased gradually,about 60% of the resulting myogenic cells were positive at 4th week after induction ,they were negative for uninduced cells.hMSCs'surface antigen profiles obtained by Flow Cytometry were negative for CD31\CD34\CD45 before and after induction,but CD90 expressed higher after induction while was weak positive before induction(P.05). Apotosis index was correlated with the cultural time after induction,The apoptosis rate of induced hMSCs was remarkably higher than control group(P.01),and the variation between groups was notable(P.05),the cell cycle analysis showed that the percentages of G_0/G_1phases were reduced significantly after induction. The expresstion levels ofβ-MHC and CTNT mRNA were undetectable before induction,began to increase at 1st、4th week after induction,reached the peak at 6th week and decreased after that,the expression of Bcl-2 and Bax mRNA varied regularly after treated with 5-azacytidine. hMSCs'resting membrane potential、range and rate of depolarization were heightened gradually after being induced.

结果：hMSCs诱导前为纺锤形，诱导后第2天部分细胞即开始发生形变，呈球形或短棒状，1周后胞浆中颗粒增多，约20～30%细胞边缘呈毛刷样变化；hMSCs表面抗原CD31、CD34、CD45在诱导前后差异无统计学意义，CD90未诱导时表达呈弱阳性，诱导后明显增高(P.05)；ANP和CX43在诱导前无表达，诱导后第2周开始表达且表达随时间逐渐增强，但CX43在诱导后第5周表达量开始降低。hMSCs诱导后凋亡指数随诱导后培养时间增加，低浓度诱导组低于标准浓度诱导组，组间差异有统计学意义(P.05)，G_0/G_1期细胞比例诱导后较对照组显著减少(P.05)；β-MHC和CTNT基因分别在诱导后第1周和第4周时表达开始增强，在第6周时均达到高峰，第8周时表达开始衰减，Bcl-2、Bax基因表达呈时间依赖性变化，hMSCs经诱导后随心肌样细胞特征的表达膜静息电位、去极化幅值和去极化速率逐渐增高。

ABSTRACT It is because that the excellent acceptors for maize transgenic engineering are insufficient in our country, especially in the southwest mountain areas of china and hereditary variation regularity for the two characters such as efficiency of embryonic callus induction and number of regenerating plant (these two characters were abbreviated to the nduction efficiency and number of regenerating in the following of the paper, respectively), which hint the maize culturing capacity, is not very clear. Therefore, aiming at picking out superior acceptors, we had made systematic researches on the two characters with combing traditional quantitative-character genetic analyzing methods such as single-factor genetic mating design, diallel crossing genetic design, genetic effect analyzing method and the modern molecular locating method such as QTLs'. The main results are followed.(1) 50 superior inbred lines and about 30 crosses in our country, especially in the southwest of China were used for identifying and selecting the superior genotypes in the above two investigated characters under the same culturing condition in 2000 and 2001. There was very significant difference among the genotypes in the both characters. But the two characters were not certainly related. Some genotypes such as 18-599 and 18-599 were very good in them. For some ones such as zong 31, induction was higher than 18-599 and 18-599 in the efficiency, but it was only 1/3 to the later in regenerating number. In some genotypes such as S37, R08, R15, P138, A318, induction efficiency was just about 3% and scarcely any regenerating plants were got. On the whole, hybrids acted better than inbreeds in the both characters.(2) Two kind of inbreeds were selected as parents of the Griffing's method 1. 18-599 and 18-599 and the inbred line zong 31 are one kind because they are not only superior in the characters of maize cross breeding, such as CA, resistance to disease and the important agricultural characters, but also excellent in transformation characters as the induction and regeneration.

针对我国、特别是西南山地所需玉米转基因工程育种优良受体极为匮乏和反应玉米幼胚培养能力的2个主要性状，即玉米幼胚胚性愈伤组织诱导率和胚性愈伤组织绿苗发生数的遗传变异规律十分不清楚的实际情况，本研究从筛选玉米转基因工程所需要的优良受体入手，采用单因素遗传交配设计、双列杂交遗传交配设计、世代基因效应等传统数量性状分析方法，以及现代分子标记定位主效QTL分析方法，对玉米幼胚胚性愈伤组织诱导率和胚性愈伤组织绿苗发生数等2个性状进行了较为系统的分析研究，取得以下主要研究结果：(1)于2000年和2001年通过对我国、特别是西南地区近50份优良自交系和近30个杂交组合，在相同培养条件下，对幼胚培养胚性愈伤组织诱导率和胚性愈伤组织绿苗发生数等2个幼胚培养能力性状进行了筛选与鉴定，发现玉米不同基因型具有完全不同的幼胚培养胚性愈伤组织诱导率和胚性愈伤组织绿苗发生数，但幼胚培养胚性愈伤组织诱导率与胚性愈伤组织绿苗发生数并不具有必然的相关关系，有的基因型，如自交系18-599和18-599在胚性愈伤组织诱导率和愈伤组织绿苗发生数等2个性状都表现相当优异；有的基因型，如自交系综31，仅幼胚培养胚性愈伤组织诱导率性状表现高于19-599和18-599，但在胚性愈伤组织绿苗发生数这一性状则与它们有相当大的差距，仅为19-599和18-599的1/3左右；有的基因型，如S37、R08、R15、P138、A318等玉米自交系不仅幼胚培养胚性愈伤组织诱导率很低，平均仅在3％左右，而且胚性愈伤组织绿苗发生数表现也很差，基本上没有分化成苗。

This research take late-maturing variety adzuki bean Jihong-4 as the experiment material, After the photoperiod inducement at different leaf ages treatment, the result showed that the activity of NR under LF treatment is higher than CK treatment on the whole, and is lower than SD treatment; The content of the dissociation amino acid under these three treatments show a descend trend along with the leaf ages accretion, but the inducement increasing the content of dissociation amino acid, different leaf ages leaves show some difference, and alternating rise with SD treatment; Inducement can reduce the content of soluble sugar, and continuous inducement shows a accumulation effect; 2LF is the most sensitivity stage for soluble protein under photoperiodic inducement, From this stage, each leaf indumenta can reducing the content soluble protein, but continuous inducement reducing rising it.

本研究通过光周期诱导的方法，对中晚熟品种冀红4号红小豆进行不同叶龄进行处理，结果表明：所有叶龄处理下硝酸还原酶活性基本高于自然光处理而低于连续短日照处理下的；随着叶龄增大，各处理游离氨基酸总量均呈下降趋势，但光周期诱导可提高游离氨基酸含量，不同叶龄表现有所差异，与连续诱导交替上升；诱导可降低可溶性糖的含量，并连续诱导出现累积效应；2LF是光周期诱对可溶性蛋变化最敏感的时期，这个时期起单个叶龄诱导可降低可溶性蛋白含量，但连续诱导却可提高；LF处理下的类胡萝素总体含量高于另两个处理，而LF处理下的叶绿素含量在仅3叶龄前高于CK处理下的，随后出现下降趋势。

The summary results are below:1. GUS expression under the driving of the BjCHI1 promoter (-1060/+17) was essentially undetectable in the young seedlings under normal growth conditions. GUS activity was first detected in the stigma of young flowers, peaked in the young siliques, and decreased when the siliques became older. No GUS expression was found in the mature siliques, seeds or root.2. The BjCHI1 promoter (-1060/+17) was inducible by NaCl, PEG, wounding and MeJA treatments. High levels of GUS expression were detected in the transgenic tobacco and Arabidopsis plants after wounding, NaCl, PEG, and MeJA treatment, indicating that the BjCHI1 promoter responses to both biotic and abiotic stresses.3. RT-PCR analysis confirmed that the expression of the BjCHI1 gene in B. juncea was inducible by PEG and NaCl.4. The transcription start site was determined by 5′-RACE, and was located at the 17th nucleotide upstream of the translation initiation codon of the BjCHI1 gene.5. A -805/+17 promoter fragment was enough to response to wounding and MeJA induction, which was proved in transgenic tobacco and Arabidopsis plants. The 397 bp region between -805 and -409 of the BjCHI1 promoter contains a cis-acting element that is essential for the wounding and MeJA inducibility.6. The -695/-620 region was necessary but not sufficient to confer MeJA-responsive expression. A T/G-box locates in -353 play an important role in the expression of the BjCHI1 gene in response to MeJA treatment. The 76 bp region is coupled with the T/G-box to confer full MeJA-inducible transcription of the BjCHI1 gene.

主要结果如下：1、利用转基因拟南芥植株分析表明，正常生长条件下，BjCHI1启动子(-1060/+17)驱动GUS基因主要在花柱中表达，幼嫩的荚也有表达，并随着果荚的成熟而减弱，成熟的果荚、种子和根没有显示GUS活性。2、BjCHI1启动子(-1060/+17)能驱动GUS基因在转基因烟草和拟南芥中响应伤害的诱导，转基因拟南芥的分析还证明BjCHI1启动子也受MeJA、NaCl和PEG的诱导，证明BjCHI1启动子是一个伤害、MeJA、NaCl和PEG等生物和非生物因素诱导启动子。3、RT-PCR进一步证明芥菜中BjCHI1基因也受NaCl和PEG的诱导表达。4、5′-RACE法鉴定了BjCHI1启动子的转录起始位点，位于翻译起始位点ATG上游第17个碱基A.5、转基因烟草和拟南芥分析证明，-805/+17的启动子片段足以响应伤害和MeJA的诱导，-805和-409之间397 bp的启动子片段含有对伤害和MeJA诱导必要的元件。6、本明烟叶片瞬时表达系统分析证明，一段76 bp的序列(-695/-620)对BjCHI1启动子响应MeJA的诱导是必要的，但不足以响应MeJA的诱导，位于-353的T/G-box也参与MeJA的诱导。76 bp的序列(-695/-620)与T/G-box协同起作用，赋予BjCHI1启动子MeJA诱导性。

The absorption and distribution of chromium were studied in ryeusing nutrient culture technique and pot experiment.

采用不同浓度K2CrO4(0，0.4，0.8和1.2 mmol/L)的Hoagland营养液处理黑麦幼苗，测定铬在黑麦体内的亚细胞分布、铬化学形态及不同部位的积累。

By analyzing theory foundation of mathematical morphology in the digital image processing, researching morphology arithmetic of the binary Image, discussing two basic forms for the least structure element: dilation and erosion.

通过分析数学形态学在图像中的理论基础，研究二值图像的形态分析算法，探讨最小结构元素的两种基本形态：膨胀和腐蚀；分析了数学形态学复杂算法的基本原理，把数学形态学的部分并行处理理念引入到家实际应用中。