低蛋白饲粮对山羊肝脏转录组的影响

doi:10.13560/j.cnki.biotech.bull.1985.2020-1505

摘要/Abstract

摘要：

为进一步实施低蛋白饲粮饲喂技术,探索低蛋白饲粮对山羊肝脏功能的调控机制。利用RNA-Seq技术对饲喂正常、低蛋白饲粮安徽白山羊肝脏组织转录组分析;利用GO富集分析和KEGG通路注释对差异表达基因功能进行解析并进行差异基因的PPI互作分析。结果表明,相比对照组,低蛋白组山羊肝脏组织中基因表达量变化在2倍以上的基因有62个（FDR<0.05）,其中,上调表达基因49个,下调表达基因13个,部分基因与蛋白质代谢、能量及糖脂代谢相关;GO富集分类结果显示差异基因共富集至42个条目,主要集中在结合、催化活性以及分子转运活性等功能;KEGG富集分析显示差异基因显著（P<0.05）富集到23条代谢通路,主要包括味觉传导、脂肪酸生物合成以及胰岛素信号通路等;PPI蛋白质互作分析发现,PPP1R3B,FASN和IRS2三个核心位置差异基因富集到胰岛素信号通路。上述结果表明,低蛋白饲粮可显著影响肉羊肝脏糖脂代谢,促进肝脏脂肪酸的合成和脂质过氧化,降低肝脏糖异生,为低蛋白饲粮的营养调控提供理论依据。

关键词: 肝脏, 转录组, 低蛋白日粮, 山羊

Abstract:

In order to further apply the low protein diet（LCP）feeding technology and investigate the regulation mechanisms of LCP on goat liver function,RNA-Seq technology was employed to obtain the transcriptome of the liver tissue of Anhui white goat fed LCP or normal diet. Go annotation and KEGG analysis were used to determine the functions of the differentially expressed genes（DEGs）,and PPI was employed for the analysis of DEGs interactions. Results showed that there were 62 genes（FDR<0.05）with over two-fold expression variation of genes in the goat liver after LCP feeding,among which 49 genes were up-regulated and 13 genes were down-regulated. A series of DEGs related to protein metabolism,energy and lipid metabolism were identified. The functions of these DEGs were classified into 42 GO items,and the DEGs were mainly concentrated in binding,catalytic activity,molecular transducer activity and other functions. KEGG analysis showed that 23 metabolic pathways were significantly（P<0.05）enriched,including taste transduction,fatty acid biosynthesis,insulin signaling pathway,and so on. Three core DEGs including PPP1R3B,FASN,and IRS2 were enriched in insulin signal pathway in the PPI network. Above results indicate that LCP significantly affects the metabolism of glucose and lipid in the goat liver,increase fatty acid synthesis and lipid peroxidation,and decrease hepatic gluconeogenesis,which provides theoretical basis for the nutrition regulation of low protein diet.

Key words: liver, transcriptomics, low crude protein diet, goat

朱雯, 汤莹莹, 孙昕旸, 周明, 张子军, 陈兴勇. 低蛋白饲粮对山羊肝脏转录组的影响[J]. 生物技术通报, 2021, 37(9): 203-211.

ZHU Wen, TANG Ying-ying, SUN Xin-yang, ZHOU Ming, ZHANG Zi-jun, CHEN Xing-yong. Effect of Low Crude Protein Diet on the Liver Transcriptome Sequencing of Goats[J]. Biotechnology Bulletin, 2021, 37(9): 203-211.

图/表 13

参考文献 24

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项目 Item	对照组CK	低蛋白组LCP
原料 Ingredient
玉米 Ground corn grain	13.0	21.0
豆粕 Soybean meal	24.0	16.0
麸皮 Wheat bran	7.5	7.5
小苏打 Sodium bicarbonate Salt	1.0	1.0
食盐Salt	1.0	1.0
磷酸氢钙 Dicalcium phosphate	0.5	0.5
石粉 Calcium carbonate	1.0	1.0
预混料Premix	1.0	1.0
花生秧 Peanut vine	28.0	28.0
羊草 Chinese wild rye	22.0	22.0
合计Total	100.0	100.0
营养水平Chemical Composition
有机物 Organic matter	86.3	88.3
粗蛋白质 Crude protein	14.8	12.0
中性洗涤纤维 Neutral detergent fiber	46.4	44.8
酸性洗涤纤维 Acid detergent fiber	31.8	31.2
粗脂肪 Ether extract	3.74	3.27
粗灰分 Ash	13.7	11.7
代谢能 Metabolizable energy	10.5	10.5

项目 Item	对照组CK	低蛋白组LCP
原料 Ingredient
玉米 Ground corn grain	13.0	21.0
豆粕 Soybean meal	24.0	16.0
麸皮 Wheat bran	7.5	7.5
小苏打 Sodium bicarbonate Salt	1.0	1.0
食盐Salt	1.0	1.0
磷酸氢钙 Dicalcium phosphate	0.5	0.5
石粉 Calcium carbonate	1.0	1.0
预混料Premix	1.0	1.0
花生秧 Peanut vine	28.0	28.0
羊草 Chinese wild rye	22.0	22.0
合计Total	100.0	100.0
营养水平Chemical Composition
有机物 Organic matter	86.3	88.3
粗蛋白质 Crude protein	14.8	12.0
中性洗涤纤维 Neutral detergent fiber	46.4	44.8
酸性洗涤纤维 Acid detergent fiber	31.8	31.2
粗脂肪 Ether extract	3.74	3.27
粗灰分 Ash	13.7	11.7
代谢能 Metabolizable energy	10.5	10.5

序号No.	引物名称Primer name	引物序列Primer sequence（5'-3'）	产物长度Product size/bp	退火温度Annealing temperature/℃
1	PC-F	TGCGGTCCATCCTGGTCAA	87	63.3
1	PC-R	ACGCCAGGTAGGACCAGTT	87	63.3
2	FASN-F	ACCTCGTGAAGGCTGTGACTCA	196	59.5
2	FASN-R	TGAGTCGAGGCCAAGGTCTGAA	196	59.5
3	IGF1-F	TCTTGAAGCAGGTGAAGATGCC	144	60
3	IGF1-R	ACACGAACTGGAGAGCATCC	144	60
4	IRS2-F	AAGCACCTATGCCAGCATCAAC	129	60.5
4	IRS2-R	GAGGATTGCTGAGGTCATTTAGGTC	129	60.5
5	CYP4A1-F	CTGCTCCGCTTTGAGCTACT	125	59
5	CYP4A1-R	GCTCCACAACGGAATTAGTGG	125	59
6	β-actin-F	TCGTGCGTGACATTAAAGAG	134	61.3
6	β-actin-R	ATTGCCGATAGTGATGACCT	134	61.3

序号No.	引物名称Primer name	引物序列Primer sequence（5'-3'）	产物长度Product size/bp	退火温度Annealing temperature/℃
1	PC-F	TGCGGTCCATCCTGGTCAA	87	63.3
1	PC-R	ACGCCAGGTAGGACCAGTT	87	63.3
2	FASN-F	ACCTCGTGAAGGCTGTGACTCA	196	59.5
2	FASN-R	TGAGTCGAGGCCAAGGTCTGAA	196	59.5
3	IGF1-F	TCTTGAAGCAGGTGAAGATGCC	144	60
3	IGF1-R	ACACGAACTGGAGAGCATCC	144	60
4	IRS2-F	AAGCACCTATGCCAGCATCAAC	129	60.5
4	IRS2-R	GAGGATTGCTGAGGTCATTTAGGTC	129	60.5
5	CYP4A1-F	CTGCTCCGCTTTGAGCTACT	125	59
5	CYP4A1-R	GCTCCACAACGGAATTAGTGG	125	59
6	β-actin-F	TCGTGCGTGACATTAAAGAG	134	61.3
6	β-actin-R	ATTGCCGATAGTGATGACCT	134	61.3

样品名称 Sample	原始数据 Raw data	筛选后的数据 Clean reads	总映射 Total mapped reads	多重映射 Multiple mapped reads	唯一映射 Uniquely mapped reads	Q30 Q30 proportion/%	GC含量 GC proportion/%
CK_1	58 699 326	58 606 316	56 533 208（96.53%）	2 878 815（4.92%）	53 654 393（91.61%）	92.09	53.06
CK_2	46 719 458	46 647 138	45 044 045（96.64%）	2 365 004（5.07%）	42 679 041（91.56%）	92.23	53.54
CK_3	46 920 008	46 857 532	44 876 131（95.83%）	2 253 785（4.81%）	42 622 346（91.02%）	92.32	52.88
LCP_1	58 554 166	58 465 664	56 561 055（96.83%）	3 000 129（5.14%）	53 560 926（91.69%）	92.32	52.65
LCP_2	47 060 314	46 984 954	45 257 361（96.43%）	2 532 160（5.40%）	42 725 201（91.03%）	92.49	53.79
LCP_3	62 169 938	62 063 962	59 438 635（95.88%）	4 063 987（6.56%）	55 374 648（89.32%）	91.87	53.59