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Tetraploid embryos could be produced by electrofusion at the stage of two-cell embryos, which could develop to blastcysts fellowed by fusion of cytoplasm and nucleus and cleavage in vitro.After fusion of cytoplasm, the DNA methylation levels of the fused embryos was very high as well as two-cell diploid embryos in vivo.Then the embryos was rapiddly demethylated when the nucleus begin to fuse, resulting the lowest DNA methylation levels when the nucleus fused completely.After that, the DNA methylation levels of fused embryos were gradually increased until the blastocysts stage.However, whereas an asymmetric distribution of DNA methylation was established in an vivo-derived blastocysts with a higher methylation level in the inner cell mass than in the trophectoderm, in most vitro-derived tetraploid blastocysts, we can not detect the asymmetric distribution.

结果表明:利用电融合方法制备的小鼠四倍体胚胎在体外培养体系中经历细胞质融合、细胞核融合及细胞继续分裂发育直到囊胚期的过程,在细胞质融合的时候胚胎卵裂球同体内体外培养二倍体胚胎一样,呈现高度甲基化状态;在细胞核开始融合的时候,甲基化水平急速下降,在细胞核完全融合的时候甲基化水平达到最低点;随着胚胎继续分裂,胚胎甲基化水平逐渐增加,在囊胚期甲基化水平最高;但是囊胚期四倍体胚胎内细胞团同滋养层细胞甲基化荧光信号没有差别,这与体内体外培养二倍体囊胚内细胞团细胞甲基化荧光强度高于滋养层细胞甲基化荧光强度不同。

In the normal group, electron microscopy showed normal chondrocytes were oval, and cells and cell membrane were intact. In cytoplasm, there were abundant rough endoplasmic reticulum, dictyosome, and mitochondria with intact nucleus, and even chromatin. In the model group, chondrocytes exhibited obvious pyknosis and their shape was irregular. The areolae around cells disappeared; cell organs in cytoplasm coagulated to flaps which showed electron dense and was uneasy to observe; the shape of nucleus showed irregularity, and caryotin gathered densely.

电镜下正常对照组软骨细胞呈椭圆形,细胞及胞膜完整,包质内可见丰富的粗面内质网、高尔基体、线粒体,细胞核完整,染色质分布均匀;模型对照组软骨细胞明显固缩且外形不规则,细胞周晕消失,胞浆内细胞器凝成高电子密度的片状物不易分辨,细胞核不规则,染色质浓聚,散裂于核中。

We also have observed some organs pathological changes of juvenile Jian Carp, such as pancreas acinous gland and zymogen granules of acinous gland cell disappearing, intercellular substance hyperplasia and inflammatory cells soakage, hepatatrophia, liver cell granular or vacuolar degeneration and necrosis, karyolysis or pyknosis, glycogen granules decreasing, metanephros atrophy, metanephric canaliculus epithelium granular or vacuolar degeneration and necrosis, mitochondrion swelling and mitochondrion cristae disappearing, karyolysis, distal convolutal tubule microvilli desquamating, spleen marrow cell degeneration and necrosis, intercellular substance of spleen hyperplasia, spleen atresia, blood corpuscle disappearing.

后肾土黄色、淡褐色或苍白色,肾小管上皮细胞肿胀、颗粒或水泡变性、坏死,细胞内有大量血细胞流出,线粒体肿胀,嵴结构消失,细胞核溶解,肾间质甲状腺滤泡和拟淋巴细胞增生,远曲小管微绒毛脱落、管道细胞界限不清。心脏肌纤维肿胀、颗粒或空泡变性,严重的肌纤维溶解、变细或断裂,肌纤维间水肿、炎性细胞浸润,部分心肌细胞核浓缩。脾脏暗褐色,脾髓质细胞变性、坏死,拟淋巴细胞明显减少,淋巴细胞岛少见、岛中细胞成份减少,黑素巨噬细胞中心减少、体积缩小,脾脏网状基质水肿,脾窦闭锁,血细胞减少。

The anastomosing occurred between the pleural superficial and the subpleural vascular networks on the capillary level or the precapillary level. The honeycomb vascular networks were also various in size and form at a low magnification. The venule connected with the pulmonary capillary networks directly at a high magnification. The single capillary networks were found in the alveolar septa. The longitudinal crests and dense round imprints of endothelial nuclei and the clear circular constrictions were found on the surface of the larger vessel in the lung parenchyma. The round imprints of endothelial nuclei also occurred on the subpleural venule and capillary.

胸膜面浅层的微血管与胸膜下微血管之间形成毛细血管水平或毛细血管前水平的吻合;低倍镜下肺实质内微血管网也呈现大小不等、形态各异的蜂窝状结构;高倍镜下肺泡毛细血管与微静脉之间联系紧密;在肺泡隔内毛细血管网成单层分布;肺实质内管径较大的微血管表面可以见到纵行的嵴状结构和密集的内皮细胞核的压痕,以及明显的环形缩窄;胸膜下微静脉和毛细血管表面可见圆形内皮细胞核的压痕。

Candidum root and surrounded it, and later penetrated the velamen cells.Or mycorrhizal fungus penetrated the exodermal passage cells through the velamencells and further infected the cells of cortex, and then formed pelotons in the corticalcells.

真菌AR-15由根被侵入外皮层通道细胞,在外皮层通道细胞内形成菌丝结,进一步向皮层细胞侵染,从而形成内生菌根,紧靠外皮层的皮层细胞具有多个细胞核和核仁,还发现细胞核正在进行穿壁运动。

In the normal uterus, Cytokeratins immunolabelling were detected in glandular cell, luminal epithelial cell, Vimentin immunolabelling were detected in stromal cell and endoblastic cell; CK7 immunolabelling were not detected in any tissue of the yak utenus.

研究结果显示:未妊娠时,泛角蛋白在子宫内膜腺上皮细胞、腔上皮细胞内表达,波形蛋白在子宫内膜基质细胞内表达,平滑肌肌动蛋白在子宫平滑肌和血管平滑肌内表达,牦牛子宫任何部位均不表达角蛋白7;妊娠30天左右时,泛角蛋白在子宫内膜腺上皮细胞、子宫内膜腔上皮细胞、滋养层细胞、内胚层细胞和尿囊细胞内表达,波形蛋白在子宫内膜基质细胞和内胚层细胞内表达,平滑肌肌动蛋白在子宫平滑肌和血管平滑肌内表达,角蛋白7在尿囊细胞内表达,偶尔在腔上皮细胞的细胞核边缘表达;消化法进行原代培养时,组织经胶原酶消化并通过100目和400目筛网组合可以有效地分离原代子宫内膜基质细胞和子宫内膜腺上皮细胞;分离得到的子宫内膜基质细胞活率达90%以上,并可在体外传代7次以上;分离得到的子宫内膜腺上皮细胞活率可达85%以上,并可在体外传代5次以上;RPMI1640培养基最适合子宫内膜基质细胞和子宫内膜腺上皮细胞的生长,维持子宫内膜基质细胞正常生长的FBS添加量为20%,维持子宫内膜腺上皮细胞正常生长的胎牛血清添加量为30%。

The results showed that ACP positive reactions were mainly deposited in lysosome, nucellar cell, cellular endomembrane, mitochondriurn, and endoplasmic reticulum in four tissues of the healthy shrimp. Furthermore, ACP activity was located in microvilli, and around partly lipid droplet in liver. AKP positive reactions were mainly deposited in cell membrane, nucellar cell, lysosome. endoplasmic reticulum and cellular endomembrane in four tissues.

结果显示,在健康的对虾体内,ACP依次出现在各组织中的溶酶体、细胞核、细胞内膜、线粒体、内质网以及肝脏组织中的部分脂滴周围及微绒毛中AKP依次出现在各组织中的细胞膜、细胞核、溶酶体、内质网以及肝脏组织中的脂滴周围及微绒毛中。

Several appear here as large cells with two or more nuclei that have prominent nucleoli.

图示可见有几个细胞内有2个或2个以上的细胞核形成了巨大的细胞核。

The results were:① The number of pCREB immunoreactive neuclei and total amount of pCREB in the subnuclei of rat amygdala, Dentate gyrus and Cornu Ammonis 3(CA3) were increased after FS.

在DG颗粒细胞层内,对照组的pCREB阳性细胞核数目很少,成线状散在分布;而FS后pCREB阳性细胞核的数目明显增多,成带状密布于整个DG颗粒细胞层。

The morphology and structure of reconstructed tissue was detected by microscope and scanning electron microscope.Results:(1) Compared with the control group, the cellular proportion of laminin group increased in 62 ~M phase, and decreased in Go~Gi phase significantly. As shown by the microscope, the cells of control group were in low density. The cells in mass connected tightly. The microfilament appeared reticular formation. The nucleus were the same in size. The cells of laminin group were in high density and put out so many lamellipodia, filopodia, which connected with the surrounding cells. The microfilament increased, elongated, and changed from reticulodromous to sarciniform, which reached to the pseudopods. The nucleus were different in size .(2) As shown by the inverted microscope and the cell growth curve, comparing with the controlgroup, cells of each test group increased evidently. The cellular proportion of each test group increased in S phase and G2 ~M phase, and decreased in Go~Gi phase significantly, but there was no considerable interations between LN and EGF;(3) As shown by the morphological observations, the cultured cat corneal endothelial cells formed an integrated membrane, and attached to the Descemets membrane closely, which was similar to the natural tissue. The cells connected tightly to each other, and some of them arranged in hexagon approximately.

结果:(1)层粘连蛋白组处于G_2~M期细胞比例较对照组显著提高,Go~G_1期细胞比例显著下降,提示层粘连蛋白促进内皮细胞DNA合成,及细胞分裂增殖;光镜下,对照组细胞分布成团状,细胞密度较低,细胞间连接紧密,细胞内微丝结成网状,细胞核大小一致;与对照组相比,层粘连蛋白组细胞生长旺盛,细胞密度高,向周边伸出大量板状及丝状伪足,细胞内微丝增多、拉长、集结成束,伸入伪足中,细胞核形状大小不一致;(2)倒置显微镜观察及细胞生长曲线显示,各组细胞数目随时间增加而明显增多,各实验组较对照组增生显著,EGF和LN联合应用组各时间点细胞数目最高;实验组处于S期和G_2~M期细胞数目增加,Go~G_1期细胞数目减少;提示EGF、LN单独及联合应用均可促进细胞增殖,但尚不能认为二者有交互作用;(3)倒置显微镜下,组织培养的猫角膜内皮细胞排列成密集的单层,细胞间连接紧密;组织学观察发现,培养的猫角膜内皮细胞形成完整的内皮层,贴附于脱水基质的后弹力膜上,与正常的角膜内皮组织结构相似;扫描电镜下,组织培养的猫角膜内皮细胞间紧密镶嵌排列,可见某些细胞呈近似六边形排列,细胞大小不甚一致,胞核清晰。

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