graces_insect_medium

Overview

External ontology

mediadive.medium:1514 (source link unavailable) — Grace's Insect Medium

Applications

• Microbial cultivation

Notes

Source: DSMZ | Link: https://www.dsmz.de/microorganisms/medium/pdf/DSMZ_Medium1514.pdf

Composition

flowchart LR
    CultureMech_000986["graces_insect_medium"]
    ing0_Grace_s_Insect_Medium["Grace's Insect Medium"]
    CultureMech_000986 --> ing0_Grace_s_Insect_Medium
    ing1_Fetal_bovine_serum["Fetal bovine serum"]
    CultureMech_000986 --> ing1_Fetal_bovine_serum
    ing2_Sucrose["Sucrose"]
    CultureMech_000986 --> ing2_Sucrose
    CHEBI_17992["CHEBI:17992"]
    ing2_Sucrose -.-> CHEBI_17992
    ing3_KH2PO4["KH2PO4"]
    CultureMech_000986 --> ing3_KH2PO4
    CHEBI_63036["CHEBI:63036"]
    ing3_KH2PO4 -.-> CHEBI_63036
    ing4_Na2HPO4_x_2_H2O["Na2HPO4 x 2 H2O"]
    CultureMech_000986 --> ing4_Na2HPO4_x_2_H2O
    CHEBI_91258["CHEBI:91258"]
    ing4_Na2HPO4_x_2_H2O -.-> CHEBI_91258
    ing5_Glutamic_acid["Glutamic acid"]
    CultureMech_000986 --> ing5_Glutamic_acid
    CHEBI_18237["CHEBI:18237"]
    ing5_Glutamic_acid -.-> CHEBI_18237

Ingredients (6)

Preparation

1. Medium for maintenance of SF-9 cells (ACC 125) for cultivation of Rhabdochlamydiae.
2. Filter-sterilize
3. Maintenance conditions of SF-9 cells (see also DSMZ catalogue for ACC 125):
4. SF-9 cells are cultivated at 25 - 29°C without the addition of CO2 in flasks (25 cm2) with tightly closed lids. When a confluent layer has formed, infection can be carried out.
5. Infection with Rhabdochlamydiae:
6. Exchange medium and add 500 – 1000 µl of EB stock solution (thawed quickly to 37°C). Centrifuge for 1 h onto the cell layer at 1600 rpm at 20°C.
7. Maintenance/Passage of infected insect cell cultures:
8. Infected cultures can be maintained e.g. in 25 cm2 flasks (e.g. Nunc flasks, VWR, 734- 2081) at 27°C in an incubator (no special requirements concerning CO2 or light). If the percentage of infected cells is still low (which might occur initially after inoculation of cultures with bacteria from a frozen stock) infected cultures may be maintained by weekly passage (1:5). Infection of yet uninfected cells by added bacteria or bacteria released from neighboring infected cells is facilitated by centrifugation (130 x g, 15 min) of the culture flask. In well-infected cultures host cell lysis will be visible (the bacteria cause host cell lysis within approximately 5-7 days after invasion of host cells). To maintain such cultures, bacteria have to be transferred by adding culture supernatant (approximately 0.5-1 ml) to fresh culture flasks containing uninfected insect cells (approximately semi-confluent culture) each 7-10 days.
9. Bacteria released from host cells can be stored in
10. at -80°C for several months.
11. Infected cell cultures (supplemented with 10% DMSO) are best stored in liquid nitrogen.

Target organisms

No target organisms are recorded.

Provenance