Identification and analysis of alternative splicing events related to sheep hair follicle development based on RNA sequencing technology

doi:10.3969/j.issn.1004-1524.20221409

Acta Agriculturae Zhejiangensis ›› 2023, Vol. 35 ›› Issue (9): 2056-2067.DOI: 10.3969/j.issn.1004-1524.20221409

• Animal Science • Previous Articles Next Articles

Identification and analysis of alternative splicing events related to sheep hair follicle development based on RNA sequencing technology

YUAN Xiaochun(), WANG Yifan, WANG Yayan, SUN Haoran, MENG Ke, LI Xinhai()

College of Agriculture, Ningxia University, Yinchuan 750021, China

Received:2022-09-28 Online:2023-09-25 Published:2023-10-09
Contact: LI Xinhai

Abstract

Abstract:

Alternative splicing is an important molecular mechanism that regulates protein diversity and genetic diversity in animals and plants. In order to identify and analyze the variable splicing during hair follicle development during depilation of Dorper sheep, the Illumina HiSeqTM 4000 sequencing platform was used to sequence the transcriptome of the skin tissue samples in the middle of the side of Dorper adult ewes in September 2019 (peak growth period, S1), January 2020 (telogen, wool growth stops, S2), and March 2020 (early growth period, new wool growth causes old wool to fall off, S3). Using the rMATS software to identify the variable splicing events and differential splicing genes in the samples, 128 033 variable splicing events were identified in three group samples, of which the proportion of skipped exons was the highest, accounting for 75.69%, and the proportion of retained introns was the lowest, accounting for 2.03%. A total of 3 448 significantly (P<0.05) different splicing genes were screened in S1 vs S2, S1 vs S3 and S2 vs S3. The functional enrichment analysis of GO (gene ontology) and KEGG (Kyoto Encyclopedia of Genes and Genomes) showed that differential splicing genes were significantly enriched in VEGF (vascular endothelial growth factor) signal pathway, MAPK (mitogen-activated protein kinase) signal pathway, PI3K Akt signal pathway, Rap1 (RAS-related protein 1) signal pathway, Hippo signal pathway, neurotrophic factor signal pathway and other pathways closely related to hair follicle development, indicating that differential splicing genes played an important role in hair follicle development during depilation in Dorper sheep, The key genes (SMAD2, FGFR2, BMP4, BCL2, etc.) that may be involved in the hair follicle development of sheep depilation traits were further screened out. By analyzing the AS events in the hair follicle development process of Dorper sheep during depilation, this paper reveals the relevant genetic regulation mechanism in the process of hair follicle development and the periodic cycle of wool, which provides a theoretical basis for the subsequent analysis and excavation of the function of development related genes in the process of hair follicle development and the periodic change of wool growth during sheep depilation.

Key words: sheep, alternative splicing, hair follicle development, differentially spliced gene

CLC Number:

S826

YUAN Xiaochun, WANG Yifan, WANG Yayan, SUN Haoran, MENG Ke, LI Xinhai. Identification and analysis of alternative splicing events related to sheep hair follicle development based on RNA sequencing technology[J]. Acta Agriculturae Zhejiangensis, 2023, 35(9): 2056-2067.

Figures/Tables 8

References 31

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基因Gene	正向引物Forward primer (5’→3’)	反向引物Reverse primer (5’→3’)	大小Size/bp
MT1C	CCCTCCTGCAAGAAGAGC	CACAGCAGCTGCACTTGTC	103
KRT23	TAGCAGATGGCAAGGTAAACAA	GTCAGATCATCCAGGGTCTTTC	160
S100A14	AGACCCTGATCAAGAATTTCCA	CCGAACTCCAGTTTAGAGTCGT	178
TPRXL	ATGCTCCCAGTGAAAGTACAGA	TCCATCTCACTGATAACTCAGCA	151
RPL6	GCCTCGGAAACTGTTGAGTC	GAGGCCCAGTCACAAGTAGC	173
GAPDH	TCCACGGCACAGTCAAGG	CACGCCCATCACAAACAT	239
RPL13	GTCAGGTACCACACGAAGGTT	CACCGAGATCCCAATGGTC	105

基因Gene	正向引物Forward primer (5’→3’)	反向引物Reverse primer (5’→3’)	大小Size/bp
MT1C	CCCTCCTGCAAGAAGAGC	CACAGCAGCTGCACTTGTC	103
KRT23	TAGCAGATGGCAAGGTAAACAA	GTCAGATCATCCAGGGTCTTTC	160
S100A14	AGACCCTGATCAAGAATTTCCA	CCGAACTCCAGTTTAGAGTCGT	178
TPRXL	ATGCTCCCAGTGAAAGTACAGA	TCCATCTCACTGATAACTCAGCA	151
RPL6	GCCTCGGAAACTGTTGAGTC	GAGGCCCAGTCACAAGTAGC	173
GAPDH	TCCACGGCACAGTCAAGG	CACGCCCATCACAAACAT	239
RPL13	GTCAGGTACCACACGAAGGTT	CACCGAGATCCCAATGGTC	105

样品名 Sample name	原始读段 Raw reads	干净读段 Clean reads	Q30/%	GC含量 GC content/%	唯一对比到的read数占比 Unique_mapped reads ratio/%	对比率 Total_mapped ratio/%
S1-1	80 305 380	80 151 468	92.90	48.51	85.82	93.65
S1-2	88 681 572	88 514 636	93.52	50.33	84.78	96.29
S1-3	88 639 604	88 435 920	93.76	50.48	84.18	95.94
S1-4	82 302 900	82 082 050	93.75	52.49	80.81	96.52
S1-5	95 132 446	94 861 532	93.53	50.14	83.97	95.97
S2-1	101 606 896	101 305 036	93.08	49.02	87.06	95.82
S2-2	111 482 124	111 138 096	93.53	50.04	85.72	96.13
S2-3	78 132 386	77 918 790	94.08	49.78	86.16	96.70
S2-4	88 934 292	88 662 238	93.50	51.37	81.77	94.38
S2-5	88 841 684	88 527 154	93.48	50.42	85.05	95.57
S3-1	94 155 288	93 804 848	92.47	51.43	83.88	95.15
S3-2	100 794 688	100 434 556	93.45	50.76	82.96	94.34
S3-3	79 139 424	78 899 488	94.13	51.69	80.02	95.91
S3-4	79 187 140	78 889 182	93.51	53.35	82.35	94.68
S3-5	80 678 998	80 361 468	93.72	51.84	77.34	92.41

样品名 Sample name	原始读段 Raw reads	干净读段 Clean reads	Q30/%	GC含量 GC content/%	唯一对比到的read数占比 Unique_mapped reads ratio/%	对比率 Total_mapped ratio/%
S1-1	80 305 380	80 151 468	92.90	48.51	85.82	93.65
S1-2	88 681 572	88 514 636	93.52	50.33	84.78	96.29
S1-3	88 639 604	88 435 920	93.76	50.48	84.18	95.94
S1-4	82 302 900	82 082 050	93.75	52.49	80.81	96.52
S1-5	95 132 446	94 861 532	93.53	50.14	83.97	95.97
S2-1	101 606 896	101 305 036	93.08	49.02	87.06	95.82
S2-2	111 482 124	111 138 096	93.53	50.04	85.72	96.13
S2-3	78 132 386	77 918 790	94.08	49.78	86.16	96.70
S2-4	88 934 292	88 662 238	93.50	51.37	81.77	94.38
S2-5	88 841 684	88 527 154	93.48	50.42	85.05	95.57
S3-1	94 155 288	93 804 848	92.47	51.43	83.88	95.15
S3-2	100 794 688	100 434 556	93.45	50.76	82.96	94.34
S3-3	79 139 424	78 899 488	94.13	51.69	80.02	95.91
S3-4	79 187 140	78 889 182	93.51	53.35	82.35	94.68
S3-5	80 678 998	80 361 468	93.72	51.84	77.34	92.41

Identification and analysis of alternative splicing events related to sheep hair follicle development based on RNA sequencing technology

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