Identification and Expression Analysis of C2H2 Gene Family in Cenchrus purpureus

doi:10.13560/j.cnki.biotech.bull.1985.2025-0902

Abstract

Abstract:

Objective To identify C2H2-type zinc finger protein gene family members in Cenchrus purpureus (elephant grass) and explore their regulatory roles in cold stress responses, thereby laying a foundation for elucidating the functional mechanisms of C2H2 transcription factors. Method Based on the whole-genome data of elephant grass(Cenchrus purpureus), bioinformatics methods were used to systematically identify members of the C2H2 gene family. Transcriptome data and RT-qPCR were employed to analyze the expression patterns of these C2H2 genes under cold stress, and heterologous expression in Saccharomyces cerevisiae was performed to verify the functions of key genes. Result A total of 144 CpC2H2 genes were identified in C. purpureus. Phylogenetic analysis clustered these genes into 10 subgroups. Gene structure and conserved motif analyses revealed that members within the same subgroup exhibited highly conserved gene structures and conserved motifs. Cis-acting element analysis showed that 65 CpC2H2 genes contained cold-responsive cis-regulatory elements in their promoter regions. Expression pattern analysis indicated that the CpC2H2 gene family displayed distinct tissue-specific expression, among which CpC2H2-33, CpC2H2-107, CpC2H2-108, and CpC2H2-131 were significantly upregulated under cold stress. RT-qPCR results were consistent with transcriptome data. Heterologous expression experiments in S. cerevisiae confirmed that the encoded products of these four genes significantly enhanced the cold tolerance of recombinant strains. Conclusion A total of 144 C2H2 genes are identified in C. purpureus. The members of this family show distinct tissue specificity and differential expression under cold stress. Among them, CpC2H2-33, CpC2H2-107, CpC2H2-108, and CpC2H2-131 can enhance the cold tolerance of heterologous organisms, suggesting that they may play key roles in cold stress responses in C. purpureus. This study provides valuable insights for further investigating the functions of the C2H2 gene family and candidate genes for improving cold resistance in elephant grass through molecular breeding.

Key words: Cenchrus purpureus, C2H2 gene family, cold stress, bioinformatics

YANG Dan, JIN Ya-rong, MAO Chun-li, WANG Bi-xian, ZHANG Ya-ning, YANG Zhi-yi, ZHOU Zhi-yao, YANG Rui-ming, FAN Heng-rui, HUANG Lin-kai, YAN Hai-dong. Identification and Expression Analysis of C2H2 Gene Family in Cenchrus purpureus[J]. Biotechnology Bulletin, 2026, 42(1): 251-261.

Figures/Tables 9

Table 1 Primers used for quantitative real-time PCR (qPCR)

基因名称 Gene name	orward primer （5′-3′）上游引物 F	everse primer （5′-3′）下游引物 R
CpC2H2-33	CCACAGGTGCTCCATCTGCC	CGTCCTCCTCCCACATCTTGC
CpC2H2-107	TCGTGCTGCGGCTTGCTGTC	GGCAACAACGTCCAACAAGCG
CpC2H2-108	CAGCGGTCGGAGGAGGAGAA	CGCAGACGGAGCACCTGAACT
CpC2H2-131	GGCGCACTACCCGATGTGGA	AGGGCGACGAGGACAGCAAG
CpEE1a	ACTACGAGCAAGAGTTGGAA	CAGGAGAAGGTCGGTAGGT

Table 2 Primers used for vector construction

基因名称 Gene name	orward primer （5′-3′）上游引物 F	everse primer （5′-3′）下游引物 R
CpC2H2-33	ATGTCTTCCATGGAGCTGC	CTCCTGATGACCGCGTGA
CpC2H2-107	ATGAAGCACCCGAGAGACAA	TGCTCGAGCTCTTCGCCTAG
CpC2H2-108	ATGTCTTCCATGGAGCTCCTCC	GCCAGGGCGTTCGACCTGA
CpC2H2-131	ATGGCGAAGCCGCAGGAG	AGCTTGGACCTCCATTTGTAG

Fig. 1 Chromosomal location and distribution of C2H2 genes in C. purpureus

Fig. 2 Phylogenetic tree analysis of the C2H2 gene family in C. purpureus

Fig. 3 Tissue-specific expression profiling of the C2H2 gene family in C. purpureus

Fig. 4 Quantitative expression analysis of candidate CpC2H2 genes in different tissues of C. purpureusLower letters indicate significant difference (P<0.05)

Fig. 5 Expression profiling of C2H2 genes in C. purpureus under cold stressHeat map visualization of RNA-seq data depicting log2 (fold change) values, with color gradients indicating relative gene expression levels

Fig. 6 Expression dynamics of candidate CpC2H2 genes in C. purpureus under cold stress

Fig. 7 Cold stress tolerance of yeast strains overexpressing candidate genes

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