Chloroplast Isolation from Arabidopsis Thaliana Plants

In Arabidopsis thaliana, most of the chloroplast proteins are synthesized in the cytoplasm encoded by the nuclear genome, and the chloroplast genome encodes 87 proteins. Sequence analysis of the Arabidopsis thaliana genome showed that about 14% of the products encoded by nuclear genes were located in the chloroplast. In addition, 4255 nuclear genome-encoded proteins in Arabidopsis are chloroplast proteins. The field of chloroplast protein import has made rapid progress in the discovery of translocon components over the past decade. Pea has been the model system of choice for studying chloroplast protein input, but Arabidopsis is now emerging as an alternative model system. However, the development of Arabidopsis in chloroplast import studies has been hindered by the inconsistency and low yield of Arabidopsis chloroplast isolation methods. Therefore, scientists need to develop simple, rapid and low-cost methods for the isolation of high-yield Arabidopsis chloroplasts.

Fig. 1. Isolation of intact Arabidopsis chloroplasts. (Aronsson H, et al., 2002)

Services

Arabidopsis thaliana can provide a large amount of leaf material due to its amazing fecundity, and the sequencing of the Arabidopsis thaliana genome is an additional model system more suitable for genetic analysis. For many years, Lifeasible has been working on the independent isolation of chloroplasts from different plant tissues. We provide the isolation of Arabidopsis chloroplasts from protoplasts. Furthermore, our scientists presented a simple, low-cost method to isolate large quantities of intact chloroplasts from Arabidopsis in approximately 1 hour. Chloroplasts isolated by this method have been successfully used in chloroplast biological research such as proteomic analysis and chloroplast protein import. The basic process for the isolation of Arabidopsis chloroplasts is as follows:

(1) For 10-day-old Arabidopsis cultures.

(2) Filter the homogenate through a double-layer Miracloth and repeat the homogenization.

(3) The pooled homogenates were centrifuged and the pellet resuspended in isolation buffer.

(4) Load the resuspended chloroplasts on a two-step Percoll gradient or a linear Percoll gradient to isolate chloroplasts.

(5) Chloroplast yield was determined using a hemocytometer.

(6) Verification of chloroplast integrity by microscopy.

Competitive Advantages

• A simple, fast and low-cost method.
• Easily applied to wild-type plants and different mutants.
• Suitable for different developmental stages of rosette leaves from 10-day-old seedlings to older plants.
• The isolated chloroplast fraction was highly pure and immunologically undetectable contamination from other organelles.
• Chloroplast isolation begins immediately after tissue harvest.
• Improving the efficiency of chloroplast protein introduction.

Lifeasible can meet the needs of customers on time and on budget through a wide range of chloroplast isolation strategies. Our aim is to be customer-centric and to provide the highest quality service to customers around the world. Our skilled and dedicated scientific researchers ensure that the most appropriate methods and techniques are selected for each specialized chloroplast project. Our customer service representatives are enthusiastic and trustworthy 24 hours a day, 7 days a week. If you are interested in our services, please feel free to contact us for more information or a detailed discussion.

References

1. Aronsson H, Jarvis P. (2002) A simple method for isolating import-competent Arabidopsis chloroplasts[J]. FEBS letters. 529(2-3): 215-220.
2. Kubis S E, Lilley K S, Jarvis P. (2008) Isolation and preparation of chloroplasts from Arabidopsis thaliana plants[M]//2D PAGE: Sample Preparation and Fractionation. Humana Press. 171-186.