rpc00010: Asparagus officinalis Asp-86 callus culture¶
- Two 9-cm Petri dishes, containing cells placed on semi-solid medium
- Subculture the cells to fresh medium immediately after arrival.
- Do not store the cell culture in a refrigerator and a freezer.
- Maintain aseptic conditions of the cell culture, and work in a laminar flow cabinet.
- Culture medium: LS medium, 1 µM 2,4-D, 1.2% (w/v) agar, pH 5.7 (medium no. 7)
- Culture conditions: 27°C, dark
- Subculture: 28-day intervals
Citation of cell line
When results obtained by using this cell line are published in a scientific journal, it should be cited in the following manner: “Asparagus officinalis Asp-86 cell line (rpc00010) was provided by the RIKEN BRC which is participating in the National BioResource Project of the MEXT/AMED, Japan.”
Asp-86 cell line was established from Asparagus officinalis L. (Terada et al. 1995 ). The Asp-86 cells are grown on a Linsmaier and Skoog (LS) medium supplemented with 1 µM 2,4-dichlorophenoxyacetic acid (2,4-D) and solidified with 1.2% (w/v) agar, pH 5.7. Our Asp-86 cell culture has been maintained in the dark at 27°C and subcultured at 28-day intervals.
Chemicals and stock solutions¶
(All stock solutions are stored at 4°C)
MS salt mix
Murashige and Skoog Plant Salt Mixture, Wako Pure Chemical Industries (#392-00591)
Chemical Concentration (mg/mL) Thiamine·HCl 0.16 myo-Inositol 40
2,4-D (1 mM)
Chemical Concentration (mg/mL) 2,4-D sodium monohydrate 0.261
(2,4-Dichlorophenoxy)acetic acid sodium salt monohydrate, Sigma-Aldrich (D6679)
KOH (1 N)
Glassware and equipment¶
- Erlenmeyer flask (200 mL), capped with two layers of aluminum foil
- Forceps, sterilized before use
Preparation of LS medium (medium no. 7)¶
Dissolve the following chemicals in approximately 800 mL of distilled water.
Chemical Amount MS salt mix 1 bag (1 L) Sucrose 30 g
Add following stock solutions, and fill up to approximately 950 mL with distilled water.
Stock solution Volume (mL) LS_VT 2.5 2,4-D (1 mM) 1
Adjust the pH of the solution to 5.7 with KOH (1 N), and fill up to 1 L with distilled water.
Pour the medium into a 200-mL flask containing 1.2% (w/v) agar.
Autoclave the flask at 121°C for 20 min.
- Pick up an appropriate amount of callus cells from a 28-day-old culture with a forceps and place the cells onto fresh LS medium.
- Incubate cell cultures under the dark condition at 27°C.
- We send Asp-86 cells on semi-solid LS medium in a 9-cm plastic Petri dish. The cells should be subcultured to fresh LS medium immediately after arrival.
- In order to maintain Asp-86 callus culture stably, it is essential to observe the growth of cells carefully. Because proliferation of Asp-86 cells is affected by culture conditions, such as a room temperature, aeration conditions of the culture and so on, an amount of cells transferred to fresh medium and the subculture intervals may vary from one lab to another.