Laboratory PI

Ruth Welti

Name of operator

Danny Vu

Date of extraction

-

Order of extraction

-

Extraction buffer

-

Temperature of extraction

75 °C for 15 min and then room temperature

Provide detailed protocol of extraction proce

from PMID: 25200898 Harvested leaves number 6, 7, and 8 were immediately dropped into 4 ml of isopropanol with 0.01% butylated hydroxytoluene (BHT) at 75?C in a 20-ml EPA vial with Teflon-lined screw-cap (Thermo Fisher Scientific, Inc., thermofisher.com). Vials were kept at 75?C for 15 min to deactivate lipid-hydrolyzing enzymes. Vials were cooled to room temperature and stored overnight at -80?C before adding 12 ml of chloroform: methanol: 300 mM ammonium acetate in water (30:41.5:3.5, v/v/v) and shaking at 100 rpm on an orbital shaker for 24 h at room temperature. Extracted leaves were transferred to a new vial and dried overnight at 105?C. The extract was stored at -80?C. Dried extracted leaves were cooled and weighed on a balance with 2 ?g detection limit (Mettler Toledo, mt.com). The precision and accuracy of the balance were previously described (Vu et al., 2012).

Where extracts stored prior to analysis

Yes

If yes, for how long?

Overnight

Conditions for storage

-80 °C

Reference for protocol

Vu et al (2014), Plant Journal