OBJECTIVETo study the tissue culture and rapid-proliferation techniques of Pueraria mirifica.

METHODThe tender branch were used as explants and cultivated in different media. The optimum media for inducing buds, proliferation and rooting were selected by adjusting the kinds and doses of plant hormones and special compounds.

RESULTThe medium of MS + IBA 0.05 mg L(-1) + BA 0.5 mg L(-1) was suitable for buds inducing and could be used in the first generation cultivation; MS + IBA 0. 02 mg L(-1) + BA 0.2 mg L(-1) and MS +BA 0.1 mg L(-1) were employed by turns in subculture, 25 days propagation coefficient was 3.0; and the medium of 1/2MS + IBA 0.1 mg L(-1) + IAA 0.2 mg L(-1) + C (special compound) 10 mg L(-1) was used for roots inducing, the rooting rate was 76.9%. Rooting plantlets were transplanted in spring and summer; the surviving rate was 81.0%.

CONCLUSIONThis technique system could be employed for rapid propagation of P. mirifica.

Pueraria ; growth & development ; Tissue Culture Techniques ; methods

In this paper we studied cryopreservation of Cyclamen persicum Mill. callus to avoid variations produced by sub-culture. The callus in the logarithmic phase after sub-culture were used for experiments. Firstly, the callus were pre-cultured in culture-medium containing 4%, 6% or 8% sucrose for different time periods, transferred to different cryoprotectants to directly cryopreserve or incubated for 2 hours at -20 degrees C, then submersed in liquid nitrogen, lastly thawed rapidly in a waterbath at 37 degrees C, and washed with liquid culture-medium containing the corresponding concentration of sucrose. Cell survival rate was computed after stained by Neutral Red, and SPSS 13.0 software was used for statistical analysis. The results showed that sucrose concentration, pre-culture time, cryoprotectants had various impacts on cell survival rate. We have developed a simple but effective protocol for the cryopreservation of callus of C. persicum. Of the different protocols tested, 4%sucrose, pre-culturing for 3 days, No. 9 cryoprotectant and freezing directly after 30 minutes at 0 degrees C results in the highest cell survival rate.
Cryopreservation ; Culture Techniques ; methods ; Cyclamen ; growth & development

As the cell co-culture techniques can better imitate an in vivo environment, it is helpful in observing the interactions among cells and between cells and the culture environment, exploring the effect mechanisms of drugs and their possible targets and filling the gaps between the mono-layer cell culture and the whole animal experiments. In recently years, they has attracted much more attention from the medical sector, and thus becoming one of research hotspots in drug research and development and bio-pharmaceutical fields. The cell co-culture techniques, including direct and indirect methods, are mainly used for studying pathological basis, new-type treatment methods and drug activity screening. Existing cell co-culture techniques are used for more pharmacological studies on single drug and less studies on interaction of combined drugs, such as collaborative compatibility and attenuation and synergistic effect among traditional Chinese medicines (TCMs). In line with the action characteristics of multi-component and multi-target, the cell co-culture techniques provide certain reference value for future studies on the effect and mechanism of combined TCMs on organisms as well as new methods for studies on TCMs and their compounds. This essay summarizes cell co-culture methods and their application and look into the future of their application in studies on TCMs and compounds.
Animals ; Biomedical Research ; methods ; Cell Culture Techniques ; methods ; Coculture Techniques ; methods ; Humans

Arachidonic acid, as an important polyunsaturated fatty acid, is identified as potential food additives or pharmaceuticals for their biological activities. In recent years, arachidonic acid production by Mortierella alpina is becoming a research highlight. The prophase relevant researches focused on the mutagenic breeding and fermentation optimization. With the depth of investigation, the advancement concerning pathway for the biosynthesis of arachidonic acid in Mortierella alpina has been made. In this review, we summarized the prophase work briefly. Mainly, we discussed the biosynthesis pathway of arachidonic acid, the key enzymes, the construction of transformation system and the genetic modification. In addition, the prospect of microorganism arachidonic acid production is put forward.
Arachidonic Acid ; biosynthesis ; Culture Media ; Culture Techniques ; methods ; Genetic Engineering ; methods ; Mortierella ; genetics ; metabolism

OBJECTIVETo acquire lots of cell to culture during the primary cell culture.

METHODWe take the split-thickness skin slide technique to acquire the dissociated fibroblast cell in two big-ear rats.

RESULTSThe cell number is above 10(6) from 1 cm x 2 cm split-thickness skin slide and the technique is simple, economic, effectve.

CONCLUSIONWe think this way is better than other methods, and should be adopted in the primary cell culture, especially in fibroblast transplantation by injection.

Animals ; Cell Culture Techniques ; methods ; Fibroblasts ; cytology ; transplantation ; Rabbits

OBJECTIVE: This study optimizes three-dimensional (3D) culture conditions of HepG2 using response surface methodology (RSM) based on the VitroGel system to facilitate the cell model in vitro for liver tissues. METHOD: HepG2 cell was 3D cultured on the VitroGel system. Cell viability was detected using Cell Counting Kit-8 (CCK-8) assay of HepG2 lived cell numbers. The proliferation of HepG2 cell and clustering performance was measured via fluorescence staining test. Albumin concentration in the culture medium supernatant as an index of HepG2 cell biological function was measured with ELISA kit. Independent factor tests were conducted with three key factors: inoculated cell concentration, cultured time, and dilution degree of the hydrogel. The preliminary results of independent factor tests were used to determine the levels of factors for RSM. RESULT: The selected optimal culture conditions are as follows: concentration of inoculated cells was 4.44 × 10 ⁵/mL, culture time was 4.86 days, and hydrogel dilution degree was 1:2.23. The result shows that under optimal conditions, the predicted optical density (OD) value of cell viability was 3.10 and measured 2.978 with a relative error of 3.94%. CONCLUSION This study serves as a reference for the 3D HepG2 culture and constructs liver tissues in vitro. Additionally, it provides the foundation for repeated dose high-throughput toxicity studies and other scientific research work.
Albumins ; Cell Culture Techniques/methods* ; Hep G2 Cells ; Humans ; Hydrogels

In comparison with their in vivo counterparts, the in vitro produced mammalian embryos had markedly lower rates of morula/blastocyst development and pregnancy after transfer to the recipients. Things became even worse in the cloned embryos. This necessitates improvement of the embryo culture system. Co-culture of embryos with different types of somatic cells was found beneficial for embryo development in vitro and many studies have been conducted in this area in recent years. In this paper, recent developments and the authors' own work in studies of co-culture of early mammalian embryos with somatic cells were reviewed, with emphasis on the effects of cell type, stage of estrous cycle and number of passages of somatic cells and supplement of serum on embryo development, and the mechanisms by which co-culture promote embryo development. The recent developments are summarized as follows: 1. Somatic cells of both homogeneous and heterogeneous origins can be used for co-culture of mammalian embryos, with similar developmental rates. 2. Supplementation of animal serum at appropriate concentrations improved the somatic cell growth and consequently the development of embryos in co-culture. 3. The estrous cycle stages of oviduct epithelial cells used for co-culture had no effect on the development of embryos. 4. Over-passaging of somatic cells reduced their efficiency in promoting development of the co-cultured embryos. In conclusion, studies have shown that co-culture overcame the block of embryo development in vitro and improved embryo quality with increased rates of implantation and pregnancy, but many problems remain to be solved on its influencing factors and mechanisms of action.
Animals ; Coculture Techniques ; methods ; Embryo Culture Techniques ; methods ; Embryo, Mammalian ; physiology ; Humans

In this article, a cell culture microchip was fabricated on the SU-8 mold based on polymer-MEMS process. In the microchip, the cell culture area was separated with microchannel by a microgap, which kept the cell culture area independent, but also regulated the micro-environment of extracellular matrix by the microfluidic flow. The cell culture microchip provided a new platform for cell research.
3T3 Cells ; Animals ; Cell Culture Techniques ; methods ; Mice ; Microfluidic Analytical Techniques ; methods

OBJECTIVETo establish and optimize the rapid propagation system of Polygonum multiflorum, as well as explore method for induction and identification of autotetraploid.

METHODPropagation medium was optimized by orthogonal test. The buds were immersed in colchicine solution with different concentrations for different time to select induction conditions for autotetraploid of P. multiflorum.

RESULTThe most appropriate propagation medium was MS medium supplemented with 1.0 mg x L(-1) 6-BA, 0.3 mg x L(-1) NAA, and 0.4 mg x L(-1) PP333. That the buds were soaked in 0.2% colchicine solution for 30 h, or soaked in 0.3% colchicine solution for 18 h, was optimal condition to induce autopolyploid of P. multiflorum with induction rate as high as 16.7%.

CONCLUSIONRapid propagation of P. multiflorum could be achieved by tissue culture. Furthermore, colchicine was an effective inducer of polyploidy, and 25 tetraploid lines were obtained through chromosome identification. The experiment laid a foundation for the wild resource conservation, superior varieties breeding of P. multiflorum.

Chromosomes, Plant ; genetics ; Culture Media ; metabolism ; Polygonum ; genetics ; growth & development ; metabolism ; Tetraploidy ; Tissue Culture Techniques ; methods

OBJECTIVEOur research studied the fast-breeding technology of Ardisia crenata sims by using tissue culture and provided the scientific foundation for industry production.

METHODThe effects of axillary buds and plant regeneration of different basic medium, hormones and additives on induction and multiplication were studied.

RESULTThe best culture medium for the induction of axillary buds, which took the stems of A. crenate were as explants, was MS + 6-BA 0.5 mg x L(-1) + NAA 0.1 mg x L(-1), and the best medium for multiplication was MS + 6-BA 2.0 mg x L(-1) + NAA 0.1 mg x L(-1) + KT 0.5 mg x L(-1), the best medium for roots generation was 1/2MS + IBA 0.2 mg x L(-1). We also found that the roots'generation, roots rate and mean number of roots can be promoted by adding 0.2% Ac, and the most suitable ground substance was river sand-perlite-vermiculite (1:1:1) or perlite-vermiculite (1:1). With axillary buds and plant regeneration methode, more than 80% A. crenata sims could be regenerated integratedly.

CONCLUSIONA. crenata sims can be regenerated integratedly and breeded fast by using axillary bud proliferation technology.

Ardisia ; growth & development ; physiology ; Culture Media ; metabolism ; Regeneration ; Tissue Culture Techniques ; methods