Effect of Lactation on Proteome of Milk Fat Globule Membrane of Laoshan Dairy Goat
We have investigated MFGM proteome of goat milk at 1, 3, 30, 90, 150, and 240 d using data-independent acquisition and data-dependent acquisition quantitative proteomics approaches. The clustering and principal component analysis revealed that the protein components in goat milk was associated with specific lactation stages. Protein-protein interaction showed that albumin had more interaction with other proteins and was considered as a central node. Our results can provide a better understanding of the MFGM proteome during the lactation cycle of dairy goats and indicate changed proteins are important for the better understanding of intrinsic physiological function of goats.
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DDA and DIA analyses were both performed on an EASY-nLC 1000 system coupled with Orbitrap Fusion Lumos (Thermo Fisher Scientific, Milford, MA, USA). The samples were loaded onto an Aqua C18 trapping column (100 μm × 2 cm, 5 μm; Thermo Fisher Scientific) with solution A (0.1% FA) at a flow rate of 300 nL/min and separated using an Aqua C18 analytical column (100 mm × 75 μm, 3 μm; Thermo Fisher Scientific). Gradient separation was performed by increasing the percentage of mobile phase B from 0% to 10% within 3 min, from 10% to 35% within 45 min, from 35% to 80% within 26 min, and to 100% in 1 min; final holding was performed at 100% for the final 15 min. For DDA analysis, mass spectrometry parameters were set as follows: MS scan range, 300−1800 m/z; normalized collision energy (NCE), 27 eV; MS resolution, 60,000; maximal injection time (MIT), 50 ms; RF lens, 30%; dynamic exclusion duration, 40 s; MS/MS high-energy collision dissociation (HCD) scan resolution, 15,000; MIT, 50 ms; and automated gain control (AGC) target, 50,000. For DIA analysis, the same nano-LC system and gradient as in the DDA analysis were used. The DIA MS parameters were set as follows: MS scan range, 395−1205 m/z; NCE, 30; MS resolution, 60,000; MIT, 100 ms; MS/MS HCD scan resolution, 15,000; and AGC target, 1e6.