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li, w.

Publications and source records attributed to li, w..

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Effect of Dietary Fat on the Metabolism of Energy and Nitrogen, Serum Parameters, Rumen Fermentation, and Microbiota in twin Hu Male Lambs

BackgroundFat is the main substance that provides energy to animals. However, the use of fat in twin Hu lambs has not been investigated. Thirty pairs of male twin lambs were examined to investigate the effects of dietary fat on the metabolism of energy and nitrogen, ruminal fermentation, and microbial communities. The twins are randomly allotted to two groups (high fat: HF, normal fat: NF). Two diets of equal protein and different fat levels. The metabolism test was made at 50-60 days of age. Nine pairs of twin lambs are slaughtered randomly, and the rumen fluid is collected at 60 days of age.\n\nResultsThe initial body weight (BW) in the HF group did not differ from that of NF group (P > 0.05), but the final BW was tended to higher than that of NF group (0.05 < P < 0.1). The digestive energy (DE), metabolism energy (ME), DE/ME in the HF group tend to be higher than those in the NF group (0.05 < P < 0.1). Ruminal ammonia nitrogen (NH3-N) and the proportion of total volatile fatty acids (TVFA) are higher than that in the NF group (P < 0.05). A high throughput sequencing analysis reveals that there were no differences between the two groups in terms of the richness estimates and diversity indices (P > 0.05). The Proteobacteria and Fibrobacteres phyla were higher than that in NF group (P<0.05).\n\nConclusionsThis study demonstrated that high fat diet before weaning can affect the abundance of several groups of rumen bacteria in rumen, such as significantly increasing phyla Proteobacteria and Fibrobacteres, and genera of Succinivibrio, Alloprevotella, and Saccharofermentans, but significantly decreasing genera of Clostridium IV, Dialister, Roseburia, and Butyrivibrio. And high fat diet improved the performance of lambs at weight gain, energy utilization, and had effect on VFA composition but no effects on serum enzymes and serum hormone.

microbiology

Direct preparation of Cas9 ribonucleoprotein from E. coli for PCR-free seamless DNA assembly

CRISPR-Cas9 is a versatile and powerful genome engineering tool. Recently, Cas9 ribonucleoprotein (RNP) complexes have been used as promising biological tools with plenty of in vivo and in vitro applications, but there are by far no efficient methods to produce Cas9 RNP at large scale and low cost. Here, we describe a simple and effective approach for direct preparation of Cas9 RNP from E. coli by co-expressing Cas9 and target specific single guided RNAs. The purified RNP showed in vivo genome editing ability, as well as in vitro endonuclease activity that combines with an unexpected superior stability to enable routine uses in molecular cloning instead of restriction enzymes. We further develop a RNP-based PCR-free method termed Cas-Brick in a one-step or cyclic way for seamless assembly of multiple DNA fragments with high fidelity up to 99%. Altogether, our findings provide a general strategy to prepare Cas9 RNP and supply a convenient and cost-effective DNA assembly method as an invaluable addition to synthetic biological toolboxes.

synthetic biology