The giant panda (Ailurapoda melanoleuca) somatic nucleus can dedifferentiate in rabbit ooplasm and support early development of the reconstructed egg

The giant panda skeletal muscle cells, uterus epithelial cells and mammary gland cells from an adult individual were cultured and used as nucleus donor for the construction of interspecies embryos by transferring them into enucleated rabbit eggs. All the three kinds of somatic cells were able to reprogram in rabbit ooplasm and support early embryo development, of which mammary gland cells were proven to be the Lest, followed by uterus epithelial cells and skeletal muscle cells. The experiments showed that direct injection of mammary gland cell into enucleated rabbit ooplasm, combined with in vivo development in ligated rabbit oviduct, achieved higher blastocyst development than in vitro culture after the somatic cell was injected into the perivitelline space and fused with the enucleated egg by electrical stimulation. The chromosome analysis demonstrated that the genetic materials in reconstructed blastocyst cells were the same as that in panda somatic cells. In addition, giant panda mitochondrial DNA (mtDNA) was shown to exist in the interspecies reconstructed blastocyst. The data suggest that (i) the ability of ooplasm to dedifferentiate somatic cells is not species-specific; (ii) there is compatibility between interspecies somatic nucleus and ooplasm during early development of the reconstructed egg.

Somatic mutations of mitochondrial genome in early stage breast cancer

The complete mitochondrial genomes of the primary cancerous, matched paracancerous normal and distant normal tissues from 10 early-stage breast cancer patients were analyzed in this study, with special attempt (i) to investigate whether the reported high

A comparative approach to somatic cell nuclear transfer in the rhesus monkey

BACKGROUND: Despite the potential utility of primate somatic cell nuclear transfer (SCNT) to biomedical research and to the production of autologous embryonic stem (ES) cells for cell- or tissue-based therapy, a reliable method for SCNT is not yet availab

Reprogramming following somatic cell nuclear transfer in primates is dependent upon nuclear remodeling

BACKGROUND: Somatic cell nuclear transfer (SCNT) requires cytoplast-mediated reprogramming of the donor nucleus. Cytoplast factors such as maturation promoting factor are implicated based on their involvement in nuclear envelope breakdown (NEBD) and prema

Impairments in embryonic genome activation in rhesus monkey somatic cell nuclear transfer embryos

Somatic cell nuclear transfer (SCNT) is a remarkable process in which a somatic cell nucleus is acted upon by the ooplasm via mechanisms that today remain unknown. Here we show the developmental competence (% blastocyst) of embryos derived from SCNT (21%)

Ontogeny of IgM-producing cells in the mandarin fish Siniperca chuatsi identified by in situ hybridisation

The ontogeny of IgM-producing cells was studied in juvenile mandarin fish Simperca chuatsi, an important fish in China's aquaculture sector. The IgM-producing cells were localised through in situ hybridisation with a probe complementary to the Ig mu-chain in lymphoid-related tissues, including head kidney, spleen, thymus, intestine and gills. In head kidney, transcripts of Ig mu were first detected at 20 days post-hatching (dph) with a few positive signals. and the number of IgM-producing cells increased obviously from 39 dph onwards. At 136 dph, a large amount of positive cells were observed in the entire organ with clusters of these cells located around the blood vessels. In spleen, IgM-producing cells were found from 26 dph onwards, followed by an increase until 67 dph: clusters of positive cells were also detected around blood vessels at 102 dph. In thymus, IgM-producing cells were first observed at 39 dph; thereafter, no obvious increase was detected until 78 dph. The positive cells in thymus were distributed mainly in the outer zone of thymus. A few IgM-producing cells were still observed in thymus of 1-year-old mandarin fish. IgM-producing cells were not detected in the intestine until 87 dph, with several discrete positively stained cells distributed in the lamina propria. IgM-producing cells, scattered mainly in primary gill filaments around blood vessels, were detected in gills from 90 dph. As in other teleosts, these results indicated that the head kidney appears to be the primary organ for IgM production in mandarin fish, and IgM-producing cells exist in all organs examined in the present study, implying their lymphoid role in fish. In addition, it is suggested that vaccination after 20 dph may be much more effective in mandarin fish. (C) 2009 Elsevier B.V. All rights reserved.

Factors affecting the efficiency of somatic cell nuclear transplantation in the fish embryo

Procedures to improve somatic cell nuclear transplantation in fish were evaluated. We reported effects of nonirradiated recipient eggs, inactivated recipient eggs, different combinations between recipient eggs and donor cells, duration of serum starvation, generation number, and passage number of donor cells on developmental rates of nuclear transplant (NT) embryos. Exposure to 25,000 R of gamma-rays inactivated recipient eggs. Single nucleus of cultured, synchronized somatic cell from gynogenetic bighead carp (Aristichthys nobilis) was transplanted into nonirradiated or genetically inactivated unfertilized egg of gibel carp (Carassius auratus gibelio). There was no significant difference in developmental rate between nonirradiated and inactivated recipient eggs (27.27% vs. 25.71%, respectively). Chromosome count showed that 70.59% of NT embryos contained 48 chromosomes. It showed that most NT embryos came from donor nuclei of bighead carp, which was supported by microsatellite analysis of NT embryos. But 23.53% of NT embryos contained more than 48 chromosomes. It was presumed that those superfluous chromosomes came from nonirradiated recipient eggs. Besides, 5.88% of NT embryos were chimeras. Eggs of blunt-snout bream (Megalobrama amblycephala) and gibel carp were better recipient eggs than those of loach (Misgurnus anguillicaudatus) (25% and 18.03% vs. 8.43%). Among different duration of serum starvation, developmental rate of NT embryos from somatic nuclei of three-day serum starvation was the highest, reaching 25.71% compared to 14.14% (control), 20% (five-day), and 21.95% (seven-day). Cultured donor cells of less passage facilitated reprogramming of NT embryos than those of more passage. Recloning might improve the developmental rate of NT embryos from the differentiated donor nuclei. Developmental rate of fourth generation was the highest (54.83%) and the lowest for first generation (14.14%) compared to second generation (38.96%) and third generation (53.01%). (C) 2002 Wiley-Liss, Inc.

西洋参体细胞胚胎发生的研究

以西洋参(Panax quinquefolium L.)为材料，从胚、胚乳、多年生主根等外植体诱导筛选得到多种胚性及非胚性愈伤组织，胚来源的胚性愈伤组织经用附加1.0ppm 2,4-D的MS培养基继代保持三年后仍具旺盛的体细胞胚胎发生能力，以胚性愈伤组织建立的液体培养系统可得到大量游离的胚状体，将胚状体包埋制成的人工种子能在附加0.1ppm NAA和1.0ppm GA_3的B_s培养基上萌发形成根、茎、叶健全的再生植株，并且移栽成活。松软型根愈伤组织以过长期继代培养后，也得到了胚状体发生。比较了多种培养因素对体细胞胚胎发生和愈伤组织的影响，其中外植体来源、基本培养基的无机盐组成以及生长素是决定愈伤组织形态结构类型和体细胞胚胎发生能力的主要因素。胚乳愈伤组织、质密型根愈伤组织以及来源于胚的松软型非胚性愈伤组织在多种诱导条件下均未能发生胚状体。 从松软型的胚或根愈伤组织均容易游离大量原生质体，用悬浮培养物游离原生质体的得率更高，从早期胚状体也可酶解得到可用于培养量的原生质体。原生质体体积均很小，培养中的行为也相似，有些形成细胞壁，细胞变形，个别进行细胞分裂，形成少数细胞团，但未形成愈伤组织;有些原生质体仍保持球形，体积剧裂膨大几十倍，小液泡汇聚成大液泡，有些液泡中积累紫红色素。 用石蜡切片、半薄切片、透射电镜和扫描电镜等方法对各种愈伤组织的内部结构及外部形态进行了详细观察，发现胚状体起源于愈伤组织的表皮或表皮下层细胞，有单个发生和成丛发生两种发生方式，不同愈伤组织细胞的显微及超微结构各具特色。胚乳愈伤组织、松软型根愈作组织以及胚状体的细胞虽然都具有某些胚性细胞结构特征，例如细胞体积小、细胞核大、细胞质浓厚、细胞器丰富等，但它们并不具有现实的体细胞胚胎发生能力。发生胚状体的细胞含大量多核糖体和内质网片段，小液泡数量少且形成多泡复合体，细胞壁也由于在组织中所外的位置不同而有厚薄两种类型之分，细胞常形成胚性细胞复合体，位于愈伤组织的外围，边缘可分化为许多小细胞团。胚性细胞复合体的表面质密平整，有许多丝状物和颗粒，细胞轮廓不清，但有显著突出的小细胞，细胞表面具沟脊;而其它各型愈伤组织的表面均为球形或半球形的大型细胞，表面仅具细小的纹理、凸起或颗粒。 胚来源的胚性愈伤组织的总蛋白组成与来源相同，但形态结构截然不同的非胚性愈作组织的总蛋白组成差异显著，后者与形态结构特征相似的根本源公软型愈伤组织具有相似的蛋白质电泳带型，但多一条33KD的蛋白带;培养基中去掉2,4-D后，松软型根愈伤组织的蛋白质组成发生轻微变化。等电聚焦和SDS聚丙烯酰胺凝胶双向电泳揭示出在单向电泳扫描上呈现最高峰的17KD蛋白质在胚性愈伤组织中随等电点的不同而分离，在其它三种构软型愈伤组织中则仅集中在等电点为5.9处，为一个大而染色深的点。 同样培养基上培养的形态结构不同的愈伤组织所含元素的量不同，说明细胞对无机盐的吸收是有选择性的，不同类型的愈伤组织可积累不同的元素。培养基中的元素组成情况在愈伤组织中也有一定反映，在含钠、钾元素较多的B_5培养基上生长的愈伤组织中这两种元素的含量也较高，培养基的离子胁迫作用和细胞对离子一定程度的被动吸收会影响细胞的代谢方式，从而导致细胞类型的分化，基本培养基对愈伤组织类型的转变及体细胞胚胎发生能力的影响可能正是通过影响细胞的离子吸收、代谢平衡而实现。