应用光学成像监测肿瘤治疗及抗微生物感染的实时过程

目的:探讨光学成像技术与基因标记技术结合的非侵入性在体分子荧光成像实时监测肿瘤治疗和抗微生物感染的过程。方法:利用绿色荧光蛋白(GFP)转染人涎腺癌细胞ACC-M,建立GFP标记肿瘤皮下肿瘤生长治疗模型,并对模型进行整体荧光成像的初步研究;同时也进行表达红色荧光蛋白(DsRed2)细菌的腹部感染及治疗模型的光学成像。结果:单克隆ACC-M-GFP细胞稳定高水平表达GFP。2只成瘤裸鼠右上侧皮下肿瘤随时间推移,荧光区域面积逐渐增加,在瘤内注射后荧光区域面积略变小。注射大肠杆菌12h后感染已有扩散发生,36h时荧光扩散至整个腹部,在48h后裸鼠死亡;同时注射卡那霉素者在12h时仅少许扩散,在36h时荧光没有扩散并且荧光强度减弱,在48h后荧光基本上已探测不到,裸鼠存活。结论:光学成像可以从时间上和空间上反映肿瘤生长及细菌消长的全过程,因而这两种模型同在体光学成像的结合为研究肿瘤治疗药物和抗生素筛选提供了一种研究平台。     

Molecular genetics of human blue cone monochromacy

Blue cone monochromacy is a rare X-linked disorder of color vision characterized by the absence of both red and green cone sensitivities. In 12 of 12 families carrying this trait, alterations are observed in the red and green visual pigment gene cluster. The alterations fall into two classes. One class arose from the wild type by a two-step pathway consisting of unequal homologous recombination and point mutation. The second class arose by nonhomologous deletion of genomic DNA adjacent to the red and green pigment gene cluster. These deletions define a 579-base pair region that is located 4 kilobases upstream of the red pigment gene and 43 kilobases upstream of the nearest green pigment gene; this 579-base pair region is essential for the activity of both pigment genes.     

RNA mimics of green fluorescent protein

Green fluorescent protein (GFP) and its derivatives have transformed the use and analysis of proteins for diverse applications. Like proteins, RNA has complex roles in cellular function and is increasingly used for various applications, but a comparable approach for fluorescently tagging RNA is lacking. Here, we describe the generation of RNA aptamers that bind fluorophores resembling the fluorophore in GFP. These RNA-fluorophore complexes create a palette that spans the visible spectrum. An RNA-fluorophore complex, termed Spinach, resembles enhanced GFP and emits a green fluorescence comparable in brightness with fluorescent proteins. Spinach is markedly resistant to photobleaching, and Spinach fusion RNAs can be imaged in living cells. These RNA mimics of GFP provide an approach for genetic encoding of fluorescent RNAs.     

用显微注射法将绿色荧光蛋白基因导入金鱼受精卵中表达

采用显微注射法将绿色荧光蛋白（GFP）基因重组表达质粒导入金鱼受精卵中，以期获得能发绿色荧光的金鱼。结果显示，在注射的3批金鱼受精卵中（第一批4310粒，第二批3952粒，第三批4056粒），分别孵出鱼苗543、282和266尾，出苗率为17.02%、12.53%和13.52%，其中表达绿色荧光的金鱼分别为23、24和18尾，表达率为4.24%、8.51%和6.77%。荧光表达检测发现，从肌肉效应期就开始检测得到绿色荧光。     

光质对彩色甜椒幼苗生长及叶绿素荧光特性的影响

研究了不同光质 (白光、红光、黄光、绿光和蓝光 )对不同品种彩色甜椒幼苗生长及叶片叶绿素荧光动力学特性的影响。结果表明 ,白光和黄光培养壮苗的效果较好 ,其他光质依次为蓝光、红光和绿光。同一光质对不同品种彩色甜椒 Chla含量和 Chla/Chlb影响差异显著 ;采用不同光质处理 ,蓝光最高 ,白光和黄光次之 ,绿光下最低。光质对不同品种彩色甜椒的荧光参数有较大影响 ,在不同光质处理下 ,白光的 Fv/Fm、Fv/Fo、ΦPS 、qp最高 ,NPQ最小 ,表明白光为最有效光 ,其次分别为黄光、蓝光、红光及绿光     

用显微注射法将绿色荧光蛋白基因导入金鱼受精卵中表达

采用显微注射法将绿色荧光蛋白（GFP）基因重组表达质粒导入金鱼受精卵中，以期获得能发绿色荧光的金鱼。结果显示，在注射的3批金鱼受精卵中（第一批4310粒，第二批3952粒，第三批4056粒），分别孵出鱼苗543、282和266尾，出苗率为17.02%、12.53%和13.52%，其中表达绿色荧光的金鱼分别为23、24和18尾，表达率为4.24%、8.51%和6.77%。荧光表达检测发现，从肌肉效应期就开始检测得到绿色荧光。     

烟草丛顶病毒编码的沉默抑制子初步鉴定

 烟草丛顶病毒（Tobacco bushy top virus,TBTV）为幽影病毒属成员,能在烟草上造成严重危害。TBTV基因组编码4个ORF,其中ORF3和ORF4几乎完全重叠,可能在TBTV的致病性中发挥重要作用。分别将TBTV的ORF3和ORF4与载体pGR107连接,构建植物表达载体pGR107 ORF3,pGR107 ORF4。经农杆菌与含外源GFP基因的pGREEN208共浸润转GFP基因的本氏烟16c叶片,2～5d后（days post inoculation,dpi)在紫外灯下观察发现浸润区有明显绿色荧光,其中pGR107 ORF4在12dpi后浸润区绿色荧光区域扩大,推断ORF4编码产物可能具有局部抑制本氏烟16c对外源GFP基因进行沉默的功能;而pGR107 ORF3浸润区绿色荧光逐渐恢复红色,GFP基因发生沉默,说明ORF3编码产物不具有抑制基因沉默的功能。进一步利用real time PCR定量检测发现,pGR107 ORF4浸润15 dpi 的本氏烟叶片中GFP RNA的含量要比仅接种pGR107的对照高,比未接种的本氏烟16c稍低。推断是因为ORF4编码蛋白抑制了植物对外源GFP RNA产生的沉默,使GFP得以在本氏烟内表达。由此进一步证实ORF4编码产物是TBTV的基因沉默抑制子。     

三个烟草品种瞬时表达OsWAK1::GFP的差异

在研究水稻基因OsWAK1功能的过程中,利用荧光分子标记的方法,将OsWAK1与绿色荧光蛋白GFP构建为融合蛋白,利用注射法使表达OsWAK1::GFP融合蛋白的农杆菌浸入烟草叶片.通过荧光显微观察.根据烟草叶片表皮细胞中是否产生绿色荧光来判断融合蛋白是否表达.研究中同时利用了3个不同的烟草品种,利用同样的方法和操作程序,但是最后根据烟草叶片中绿色荧光的检测结果发现,不同的烟草品种对OswAK1::GFP融合蛋白的表达差异很大.OsWAK1::GFP融合蛋白能够在心叶烟叶片中表达,却未能检测到其在本生烟和三生烟两个品种的叶片中表达.     

种内密度效应对豌豆蚜种群增长的影响

[目的]明确红色型和绿色型豌豆蚜(Acyrthosiphon pisum)的种群数量变化机制,探索2种色型豌豆蚜的种内密度效应对其种群增长的影响。[方法]在温室内设置不同的密度梯度,单种色型豌豆蚜密度设置为8、16、24、32头/株,2种色型共存时红色型和绿色型密度分别设置为4∶4、8∶8、16∶16头/株。[结果]单种色型豌豆蚜从密度8头/株增至32头/株时,随着密度的升高,2种色型豌豆蚜的产蚜期和寿命显著缩短,产蚜量显著降低,说明2种色型豌豆蚜种群密度效应均存在一定的负效应。在总密度为8和16头/株时,2种色型豌豆蚜混合饲养的产蚜期、寿命与单种色型饲养相比无显著差异;在单种色型饲养的密度与混合饲养的同一色型密度相同时,由于混合饲养的总密度高于单种色型饲养,混合饲养的产蚜期、寿命显著短于单种色型饲养,产蚜量显著低于单种色型饲养;在密度为32头/株时,单种群饲养的产蚜期、寿命、单头产蚜量与混合饲养时无显著差异。在单种色型饲养条件下,同一密度下(16头/株除外)红色型豌豆蚜的发育历期显著长于绿色型豌豆蚜,在相同密度条件下,红色型豌豆蚜的产蚜期、寿命、产蚜量与绿色型豌豆蚜相比无显著差异;混合饲养中,在同一密度条件下,红色型豌豆蚜的产蚜期、寿命、产蚜量与绿色型豌豆蚜相比无显著差异。[结论]红色型豌豆蚜在该环境条件下的竞争力与绿色型豌豆蚜相当,红色型豌豆蚜与绿色型豌豆蚜共存对环境表现了一定的适生性。     

绿色荧光蛋白在IGF2b基因慢病毒载体感染鲤受精卵中的标记效果

以水母绿色荧光蛋白基因为模板进行PCR扩增得到目的基因(Green fluorescence protein, GFP),然后加上酶切位点BamHI和NheI,构建pLenti6.3IRESEGFP载体,转染DH5α感受态细胞进行菌落PCR,取阳性进行酶切鉴定,再取呈阳性的质粒进行测序,使用浓度为1 μg/μL的质粒与慢病毒表达载体进行连接,通过荧光显微镜观察到绿色荧光,表明本实验获得的GFP和慢病毒载体整合成功;用此转染293T细胞,通过荧光显微镜同样检测到了绿色荧光。使用建鲤的组织提取RNA,然后按照Fermentas公司的MMLV操作说明书进行反转录,得到IGF2b基因后加酶切位点进行扩增,将IGF2b整合到用GFP作为标记基因的慢病毒载体上,再以此转染建鲤未分裂的受精卵,48 h后通过荧光显微镜也观察到了绿色荧光蛋白的表达。试验表明绿色荧光蛋白在IGF2b基因慢病毒载体感染鲤受精卵中的标记是成功的。这些结果为基于含有GFP慢病毒转基因鱼育种技术的开发奠定了基础。     

Tandem array of human visual pigment genes at Xq28

Unequal crossing-over within a head-to-tail tandem array of the homologous red and green visual pigment genes has been proposed to explain the observed variation in green-pigment gene number among individuals and the prevalence of red-green fusion genes among color-blind subjects. This model was tested by probing the structure of the red and green pigment loci with long-range physical mapping techniques. The loci were found to constitute a gene array with an approximately 39-kilobase repeat length. The position of the red pigment gene at the 5' edge of the array explains its lack of variation in copy number. Restriction maps of the array in four individuals who differ in gene number are consistent with a head-to-tail configuration of the genes. These results provide physical evidence in support of the model and help to explain the high incidence of color blindness in the human population.     

Activity-dependent transfer of brain-derived neurotrophic factor to postsynaptic neurons

Neurotrophins such as brain-derived neurotrophic factor (BDNF) are thought to be transferred from post- to presynaptic neurons and to be involved in the formation and plasticity of neural circuits. However, direct evidence for a transneuronal transfer of BDNF and its relation to neuronal activity remains elusive. We simultaneously injected complementary DNAs of green fluorescent protein (GFP)-tagged BDNF and red fluorescence protein into the nucleus of single neurons and visualized expression, localization, and transport of BDNF in living neurons. Fluorescent puncta representing BDNF moved in axons in the anterograde direction, though some moved retrogradely, and transferred to postsynaptic neurons in an activity-dependent manner.     

水稻OsWrky基因的细胞核定位研究

绿色荧光蛋白（GFP）基因是一种被广泛应用的报告基因。为了了解克五笔型的水稻WRKY（OsWrky）基因的核定位性质，将OsWRKY与GFP基因融合，并构建在Ubiquitin启动子控制的pCambia130载体上（pCU-oSwrkyGFP)。利用基因枪介导的方法将pCU-OsWrkyGFP导入洋葱内表皮后，荧光显微镜观察GFP的发光部位仅在细胞核内，而作为对照的没有融合OsWRKY基因的载体（pCU-GFP)轰击后细胞质和细胞核中都有荧光。充分说明OsWRKY能作为一个转录因子在细胞核内起作用。     

中国萝卜皮色遗传的初步研究

通过对中国萝卜（ＲａｐｈａｎｕｓｓａｔｉｖｕｓＬ．ｖａｒ．ＬｏｎｇｉｐｉｎｎａｔｕｓＢａｉｌｅｙ）不同皮色试材的杂交，Ｆ１、Ｆ２自交，研究了皮色遗传表现。初步结果表明，皮部绿色（即绿皮）依不同杂交组合，可能有不同的遗传模式，即可由１对基因控制，也可由２对连锁的基因或独立分配的基因对控制。若为连锁的２对基因控制时，它们还受到细胞质基因组（可能主要是质体基因组）的互作，表现出明显的偏母遗传现象。皮部红色（即红皮）似有３对独立遗传的基因控制，这３对基因还有相互作用，其中１对可能与绿色控制基因连锁。这样，红皮与绿皮试材杂交时，参与遗传的基因可达４～５对，它们之间出现复杂的相互作用，而使Ｆ２代出现暗紫、洋红、绿色、紫绿等众多不同颜色。其中有的基因还可能影响到肉质色的遗传，使暗紫色Ｆ２代个体自交，Ｆ３代分离出少数紫皮红心萝卜，将其再自交，则能分离出绿皮红心萝卜，从而在实验过程中证实和重演了绿皮红心萝卜的起源。     

不同饲料对棉铃虫幼虫体色遗传及药剂敏感性影响的研究(英文)

不同饲料对棉铃虫幼虫体色变化有影响 ,用嫩棉叶喂养的幼虫 ,其体色以黑色、红色、绿色、绿褐色为主 ,其中黑色比率最高达 32 3% ;用人工饲料喂养的幼虫 ,其体色以绿褐色、黑色、红色、灰褐色为主 ,其中绿褐色比率最高 (31 6 % ) .利用纯化 3代的不同体色交叉组合配对 ,其后代体色遗传不规律 ,与其亲代体色相一致的幼虫占多数 .利用质量浓度为 1 0 %的功夫和 2 0 5 %的保丰乳油 ,点滴法测定触杀效果 ,结果表明 ,绿色和灰褐色是较敏感的类型 ,红色和黑色为不敏感的类型     

Motion perception: a color-contingent aftereffect

When observers who watched repeated alternations of a red contracting spiral and green expanding spiral were later shown stationary spirals, red and a green the red stationary spiral appeared to be expanding and the green stationary spiral appeared to be contracting. These color-contingent motion after effects complement reports of motion-contingent color aftereffects and suggest that both may reflect adaptation of detectors specific to color and motion.     

"Fluorescent timer": protein that changes color with time

We generated a mutant of the red fluorescent protein drFP583. The mutant (E5) changes its fluorescence from green to red over time. The rate of color conversion is independent of protein concentration and therefore can be used to trace time-dependent expression. We used in vivo labeling with E5 to measure expression from the heat shock-dependent promoter in Caenorhabditis elegans and from the Otx-2 promoter in developing Xenopus embryos. Thus, E5 is a "fluorescent timer" that can be used to monitor both activation and down-regulation of target promoters on the whole-organism scale.     

两种海棠秋季叶色变化的生理机制研究

以王族海棠(Malus‘Royalty’)和光辉海棠(Malus‘Guanghui’)秋季(8~10月)成熟叶片为试验材料,观察其叶色变化,并对其叶绿素、花色素苷、可溶性糖含量及苯丙氨酸解氨酶(PAL)活性进行测定,以期揭示两种海棠叶片色泽、内色素变化与其内源物质的相关关系。结果表明,王族海棠叶片虽生育期呈现紫红色,但秋季却出现"返绿"现象;光辉海棠则随着温度升高,叶片由绿色转变为红色,温度降低,红色退去,叶片呈现绿色。上述指标测定结果显示,两种海棠叶色表现均与叶片内色素的比例密切相关。王族海棠叶片内当花色素苷与叶绿素含量比值降至14时,其叶色由紫红色转变为绿色,其叶色变化主要是由花色素苷降解而叶绿素的积累引起的;光辉海棠成熟叶片的叶色变化则是由花色素苷和类胡萝卜素的含量共同决定的,花色素苷与叶绿素含量比值升至3、类胡萝卜素大量积累时,其叶色由绿色转变为橙红色。可溶性糖为花色素苷的合成提供原料和能源,PAL活性和叶片内色素的合成有必然联系,但不是惟一决定色素合成的调控酶。     

Effect of Light on Flavonoids Biosynthesis in Red Rice Rdh

The effect of light on flavonoids biosynthesis in red rice Rdh was studied. The panicles of red rice Rdh produced colorless caryopses after darkness treatment; and these colorless caryopses displayed bright-red after vanillin treatment, but did not display red color after light inducing for 15 days, suggesting that red rice Rdh could produce leucoanthocyanidin, but could not produce polyproanthocyanidins in darkness. Histological study revealed that the aleurone layers of Rdh colorless caryopses displayed bright-red after vanillin assay, but the pericarp and seed coat layers did not display color change, which indicated that the aleurone layers could accumulate precursors of polyproanthocyanidins in darkness, but the pericarp and seed coat could not. Additionally, color ofRdh caryopses changed from green in immaturity to red in maturity, and the green caryopses changed color from green to red gradually indoor for 7 days after harvest, suggesting that leucoanthocyanidins could synthesize polyproanthocyanidins. It was concluded that light was necessary for red pigment biosynthesis in red rice Rdh, leucoanthocyanidins biosyntheses in the aleurone layers did not need light, leucoanthocyanidins biosynthesis in pericarp and seed coat needed light inducing, the effect of leucoanthocyanidin biosynthesis in Rdh to light had tissue specificity.     

Effect of Colored Sticky Cards on Non-target Insects

Field experiments to evaluate four different colored sticky cards for trap-ping non-target insects were conducted in an organic maize field in the Heinigou region of China. Yel ow, blue, green, and red...