Fen zi xi bao sheng wu xue bao = Journal of molecular cell biology最新文献

By observing the chromosomal spreads of germ cells and tissue sections, we studied the chromosomal spreads of germ cells in diploid and polyploid fish produced by distant crossing. The samples covered the second generation of hybrids of red crucian carp (Carassius auratus red var.) (female) x common carp (Cyprinus carpio) (male) (2n = 100) (F2), allotetraploid hybrids of red crucian carp (female) x common carp (male)(4n = 200), triploid hybrids of gold fish (female)x allotetraploid (male) (3n = 150), the second generation of the gynogenetic progeny of allotetraploid hybrids (G2) (2n = 100), and the common carp (2n = 100) used as a control. The results demonstrated that chromosomal number of spermotogonia in common carp was equal to that of their somatic cell (2n = 100), while the chromosomal number of germ cells in diploid hybrid fish and polyploid hybrid fish had doubled obviously, and the frequent of chromosomal doubling in spermotogonia of F2 appeared especially high, accounting for 21.6% of all examined chromosomal spreads, which provided directly cytological evidence for the production of unreduced diploid gametes in F2 and also indicated that distant crossing was an important factor to lead to chromosomal doubling in germ cell. This research had important significance in studies on the formation of polyploidy fish and fish breeding.

Potassium antimonite precipitation was used to locate calcium in the central cell of lettuce (Lactuca sativa L.) before and after pollination. At 3d before anthesis, two polar nuclei of central cell separately located at two polarity of the cell, and few calcium precipitates (ppts) appeared in the polar nuclei and cytoplasm, but some ppts in its small vacuoles. At 2d before anthesis, two polar nuclei moved toward the middle of the cell and fused to form a secondary nucleus, and the ppts evidently increased in the nucleus and cytoplasm. At 1d before anthesis, secondary nucleus again moved toward micropylar end and located near the egg to prepare for fertilization. Calcium precipitates were mainly accumulated in the secondary nucleus. After pollination and before fertilization, the distribution of calcium ppts was similar to that before pollination. At 4h after pollination, the central cell was fertilized, and calcium ppts evidently increased in the cell and numerous were accumulated in its nucleus and cytoplasm. At 6h after pollination, the primary endosperm nucleus completed its first division and formed two dissociate endosperm nuclei, and still many calcium precipitates appeared in the nucleus and cytoplasm. With endosperm development, calcium ppts decreased in the endosperm cell. At 1d after emasculated and without pollination, the secondary nucleus of the cell still bordered on the egg and some calcium ppts appeared in the secondary nucleus. The results indicated that the temporal and spatial changes of calcium in the central cell may play an important physiological role during the development of the central cell and endosperm.

Using petioles as explants, a cotton laccase cDNA (GaLA C1) was introduced into Populus alba var. pyramidalis by A. tumefaciens-mediated transformation. PCR and Southern blot analysis indicated that transgene was stably integrated into the genome of transformants. Enzyme assay showed that laccase activity was obviously increased in transformants. As compared with untransformed control, total lignin content in all tested transgenic lines was elevated in varying degrees (as highest as 21.5%). Histochemical staining of lignin further confirmed that overexpressing GaLA C1 could result in increased lignin content in transformants. Together, our data strongly suggested that GaLA C1 may participate in lignin synthesis and this is the first direct transgenic evidence for the involvement of plant laccases in lignification.

To illustrate distribution of fat-soluble compounds in the roots, stems and leaves of four Salvia plants, the methods of Histochemistry and HPLC were adopted to analyze different parts of the four Salvia plants in this paper. The results showed that distribution was differential, and following as this: the roots, stems and leaves of four Salvia plants contained fat-soluble compounds, moreover, the fat-soluble compounds of the roots located in periderm and the stems and leaves in epidermis. The main components of the fat-soluble compounds were Tanshinone IIA, Tanshinone I and Dihydrotanshinone I in the toots of Salvia miltiorrhiza Bunge and Salvia miltiorrhiza bge. f. alba, yet there were only Tanshinone IIA in the roots of Salvia japonica and Salvia officinalis. And fat-soluble compounds were not Tanshinone IIA, Tanshinone I and Dihydrotanshinone I in the stems and leaves of four Salvia plants. The type and content of fat-soluble compounds related to the species and introduction regions, they changed with the species and introduction regions. The conclusion clarified the accurate distribution of fat-soluble compounds in the different parts of four Salvia plants, and provided some theoretical basis for the application of Chinese herbs.

We constructed a recombinant plasmid of water channel protein Aquaporin 1 (AQP1) carboxyl terminal domain (DNA sequence from 700bp-801bp) in pGEX-4T-1 vector and express the carboxyl terminal hydrophilic peptide AQP1 in E. coli. In this study, the DNA sequence of AQP1 hydrophilic peptide was amplified by PCR and was cloned into pGEX-4T-1 expression vector. After identified by restriction enzyme digestion and sequencing, the recombinant clone was transformed into the competent expression cells of E. coli BL21. The GST-AQP1 fusion protein was induced by IPTG and further purified by Glutathione Sepharose 4B to obtain a fusion protein with molecular weight of 30KD. So the fusion protein of AQP1 C-terminal hydrophilic peptide combined with GST was successfully expressed and purified. We set up important bases for the further research in AQP1 gene function.

Cloning by somatic cell nuclear transfer has been achieved by both electric fusion and intracytoplasmic nuclear injection (ICNI) methods. However, each of the above methods involves extended complicate manipulation and special equipment. Here we report a whole-cell injection technique without Piezo assistance for nuclear transfer in pigs. The fibroblast cell of pig as the nucleus donor cell, effects of the new method on the efficiency of somatic nuclear transfer in pig were investigated, compared with that of electric fusion method. Results showed that the new method was a little less efficient in producing reconstructed embryos but without significant difference (88.4% vs 78%, P > 0.05). After the embryos were cultured 48h and 7d, the fusion method is more efficient than the new method in the oocyte cleavage rate and the blastocyst development (78% vs 53.2%, P < 0.05; 27.2% vs 13.8%, P < 0.01). The results indicate that both methods make no difference in the quality of the blastula, but the electric fusion method is more efficient. Therefore, the applicability of producing normal,cloned piglets by the simple and less labor-intensive whole-cell intracytoplasmic injection needs further improvement.

Anther wall is general and tapetum is glandular. The process of meiosis of microspore mother cells is simultaneous and the tetrads are tetrahedral. The mature pollen of Pseudostellaria heterophylla (Miq.) is tree-celled. There are 22-30 germ pores on the pollen wall. Many pollen grains could burst in 10% mannitol or 15% sucrose solution and release a pair of sperm cells which could keep alive for 25-50 min by FDA fluorescence. Using micromanipulator the released sperm cells could be collected. When pollen grains were put into a solution containing 0.03% CaCl2, 0.01% H3BO3, 0.01% KH2PO4 and 20% PEG for 2-5 min, they would germinate and the pollen tubes would reach 815 microm at 2h after cultured. A pair of sperms would enter into pollen tube when it grew to 500-600 microm. The fluorescence of both sperms would be observed clearly in pollen tube after DAPI staining. When the pollen tubes were burst in a bursting solution, a pair of sperms would be released from pollen tube.

Polysaccharides was detected using the periodic-acid-Schiff's (PAS) technique and lipid detected using Sudan black during the anther development of Allium cepa L. Before the meiosis of microspore mother cells there were a few lipid drops in endothecium cells and little polysaccharides in tapetal cells which did not differentiate completely in young anthers. At the stage of tetrad there were still few polysaccharides and lipid material in young anther, and only cell wall of anther wall and callose wall of tetrads displayed the feature of polysaccharids. The size of tapetal cells began to increase at this stage. During microspore development the tapetal cells reached its maximal size, and many lipid drops were accumulated in the cells. However, few lipid drops and starches appeared in microspores. At early stage of 2-celled pollen, the vegetative cell of 2-celled pollen began to accumulate starches. Tapetal cells degenerated at this stage and its lipid drops concentrated to form lipid block. Then the starches in 2-celled pollen disappeared with pollen development, and many lipid drops were accumulated in vegetative cell of nearly mature pollen.

The expression of galanin and galanin receptor-2 in hippocampus and dorsal raphe nucleus of depression model was studied. The chronic stress rat model was adopted as the modal of depression. Open-field test was used to observe the transformation of their behavior and HPLC-EC was employed to analysis the level of blood serum cortisol. The method of in situ hybridization was used for testing the expression of Galanin and galanin receptor-2 in hippocampus and dorsal raphe nucleus and the method of RT-PCR was used to further analysis of the expression. The results showed that the locomotion activity decreased extremely after chronic stress, but the level of serum cortisol increased evidently. The expression of Galanin and its receptor-2 in hippocampus and dorsal raphe nucleus increased obviously. The higher expression for galanin and galanin receptor-2 in some brain area suggested that galanin probably takes part in the modulation of the function of neurons during the stress process.

To investigate the methylation pattern of 12 CpG dinucleotide sites in promoter region of LDLR gene in atherosclerosis patients, peripheral blood DNA samples were prepared from 61 atherosclerosis patients and 28 healthy subjects. The methylation status of CpG islands in LDLR gene promoter region was measured by using a modified coordinative method with nested-PCR, Touchdown-PCR and methylation-sensitive-single-strand conformation analysis (MS-SSCA). Three types of methylation patterns were detected in promoter region of LDLR gene in peripheral blood genome of atherosclerosis patients: full-methylation, part-methylation and none-methylation. 2 cases were full-methylation and 13 cases were part-methylation, the methylation frequency was 24.59% in atherosclerosis patients. While in 28 healthy control, only 1 half-methylation sample was found, the methylation frequency was 3.57%, which is much lower than that in atherosclerosis patients (P < 0.05). A significantly higher methylation degree in promoter region of LDLR gene was found in peripheral blood genome of atherosclerosis patients compared with the controlled healthy subjects, which may suggest an involvement of aberrant methylation of LDLR gene in initiation and development of AS. The modified coordinative method with nested-PCR, Touchdown-PCR and MS-SSCA manifests merits of convenience, cheap and high efficiency, which lowers the requirement for the DNA template and increases the sensitivity and specificity.