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发布于:2018-2-10 00:00:03  访问:1 次 回复:0 篇
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About How Ixazomib Helped Me Turning Rich And Famous
The results for between four and seven replicates for each mutant were compared to those for 14 replicates for Ler. Metabolite profiling was performed by GC-MS as previously described (Lisec et?al., 2006). Briefly, weighed powdered samples (approx 20?mg fresh weight) were extracted using chloroform:methanol:water (2/5/2 v/v/v). The extracts were then centrifuged (10?min at 11?000?g), and the supernatant was taken, dried and derivatized using methoxyamine hydrochloride in pyridine followed by N-methyl-N-trimethylsilyltrifluoroacetamide (MSTFA). Derivatized samples were used for GC-TOF analysis using an Agilent 6890 gas chromatograph (http://www.agilent.com) coupled to a LECO Pegasus?2 mass spectrophotometer (LECO, http://www.leco.com/). Twenty spectra per second were recorded selleck between m/z 85 and 500, and Pegasus software (LECO) was used to export the chromatograms as raw files in the NetCDF format. Targeted metabolite identification was performed using the TargetSearch bioconductor package (Cuadros-Inostroza et?al., 2009) with a library based on approximately 900 reference compounds from the GMD database (Kopka et?al., 2005; Schauer et?al., 2005). Only compounds with a retention index (RI) deviation of <2000 and an identification based on at least five matching correlated masses were retained. Redundant metabolites were either removed or grouped to retain the most likely identification, and other potential hits were noted (see Table?S1). Metabolites with significant differences between groups were identified based on a Kruskal�CWallis test AZD8055 with a false-discovery rate corrected P value?learn more were designed based on the polymorphisms between Ler and Col-0 described in the Monsanto Arabidopsis Polymorphism Collection database (http://www.arabidopsis.org). For allele sequencing, PCR products spanning the At1g29900 and At3g27740 transcription units were obtained using wild-type and mutant genomic DNA as templates and the oligonucleotides shown in Table?S2. The sequencing reactions were performed using an ABI PRISM BigDye Terminator cycle sequencing kit (Applied Biosystems, http://www.appliedbiosystems.com/) in 5?��l reaction volumes. Sequencing electrophoresis was performed using an ABI PRISM 3130xl genetic analyzer (Applied Biosystems). For co-expression in E.
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