Dear Muzaffer,
Below is the protocol used in our lab. Good luck.
Hormone extraction using methanol and chloroform extraction
1. weigh plant samples 20 m g FW(e.g., Arabidopsis seeds)
2. grind in 200ul 90% methanol to powder using GenoGrinder 1 min
3. add 200 ul 90% methanol, vortex
4. spin at 12000 rpm 10 min, transfer supernatant to a glass tube
5. adding 600 ul 90% methanol, vortex, combine supernatant (total 1 ml)
6. add 0.5 ml chloroform and 0.6 ml water, mix briefly
7. allow phase-separation in cold room for 1 h
8. Remove the upper aqueous phase
9. concentrate to the aqueous phase to dryness in a speedvac
10. redissolve in 80 ul water
11. spin down pellet and transfer supernatant to an HPLC vial
Follow our paper.
Cold-adapted and rhizosphere-competent strain of Rahnella sp. with broad-spectrum plant growth-promotion potential.
Pratibha Vyas, Robin Joshi, K C Sharma, Praveen Rahi, Ashu Gulati, Arvind Gulati, Journal of Microbiology and Biotechnology 12/2010; 20(12):1724-34
Dear Robin Joshi,
We study on plant hormons but your papers on bacteria strain.
Thanks.
Dear Muzaffer,
for my PhD thesis I used a HPLC protocol for ABA determination in apple samples. Please find attached a pdf of one of those papers. For more detail, please refer to the reference therein to Carrasquer et al. (1990).
I hope to be of some help.
Best wishes.
Dear ISabel Lara,
I read your manuscript but i didnt find protocols of ABA also didnt find Carrasquer et al. (1990) in this paper.
Would you like to check it for me? Please
Indeed, you are right!!! I mistook the pdf. Please find attached the correct one.
Sorry for that. I hope now it is the right one.
In my doctoral thesis I analyzed by HPLC endogenous IAA content in chestnut and oak microshoots. Please find attached the pdf files. Best wishes
Hocam ekte bir tez gönderiyorum. HPLC de GA3 ve ABA bakılmış bi ekstraksyon protokolü var yakınlarda bizde onu kullanmayı düşünüyoruz.
http://traglor.cu.edu.tr/objects/objectFile/1q1o66M6-2982013-42.pdf
Mehmet Hocam,
Tez için teşekkürler, mezuniyetimi yaptığım ve tanıdığım bir abimizin tezi olduğu için sevindim. İnşallah protokolü kullanılabiliriz.
Dear Muzaffer
You can find a protocol for ABA in my paper about ABA in orchid seeds, but it is an older method. You could also try extraction and clean-up on solid phase, if necessary followed by enrichment on HPLC and a final separation and quantification with GC/MS in SIM-mode and use of deuterated ABA as an internal standard.
Best regards
Quantitative analysis of major plant hormones in crude plant extracts by high-performance liquid chromatography–mass spectrometry
Xiangqing Pan1,2,4, Ruth Welti3 & Xuemin Wang1,2
Nature Protocols 5, - 986 - 992 (2010)
Published online: 6 May 2010 | doi:10.1038/nprot.2010.37
Dear Muzaffer,
Below is the protocol used in our lab. Good luck.
Hormone extraction using methanol and chloroform extraction
1. weigh plant samples 20 m g FW(e.g., Arabidopsis seeds)
2. grind in 200ul 90% methanol to powder using GenoGrinder 1 min
3. add 200 ul 90% methanol, vortex
4. spin at 12000 rpm 10 min, transfer supernatant to a glass tube
5. adding 600 ul 90% methanol, vortex, combine supernatant (total 1 ml)
6. add 0.5 ml chloroform and 0.6 ml water, mix briefly
7. allow phase-separation in cold room for 1 h
8. Remove the upper aqueous phase
9. concentrate to the aqueous phase to dryness in a speedvac
10. redissolve in 80 ul water
11. spin down pellet and transfer supernatant to an HPLC vial
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