Comprehensive characterization and expression analysis of enzymatic antioxidant gene families in passion fruit (Passiflora edulis).

Passion fruit, a valuable tropical fruit, faces climate-related growth challenges. Antioxidant enzymes are vital for both stress protection and growth regulation in plants. We first provided systemic analysis of enzymatic antioxidant gene families in passion fruit, identifying 90 members including 11 PeSODs, 45 PeAPXs, 8 PeCATs, 7 PeGPXs, 6 PeMDHARs, 8 PeDHARs, and 5 PeGRs. Gene members in each gene family with same subcellular localization showed closer phylogenetic relationship. Many antioxidant genes exhibited tissue- or developmental stage-specific expression patterns during floral and fruit development, with some widely expressed. Their co-expressed genes were linked to photosynthesis and energy metabolism, suggesting roles in protecting highly proliferating tissues from oxidative damage. Potential genes for enhancing temperature stress resistance were identified. The involvement of diverse regulatory factors including miRNAs, transcription factors, and CREs might contribute to the complex roles of antioxidant genes. This study informs future research on antioxidant genes and passion fruit breeding.

PeHVA22 gene family in passion fruit (Passiflora edulis): initial characterization and expression profiling diversity.

Passion fruit, an economically valuable fruit crop, is highly vulnerable to adverse climate conditions. The HVA22 genes, recognized as abscisic acid (ABA) and stress-inducible, play vital roles in stress response and growth regulation in diverse eukaryotic organisms. Here, six HVA22 genes were firstly identified in passion fruit genome and all predicted to be localized within the endoplasmic reticulum. Phylogenetic analyses showed that all PeHVA22s were divided into four subgroups. The gene structural features of PeHVA22 genes clustered in the same subgroup were relatively conserved, while the gene structure characteristics of PeHVA22s from different subgroups varied significantly. PeHVA22A and PeHVA22C closely clustered with barley HVA22 in Group II, were also induced by ABA and drought stress treatment, suggesting conserved roles similar to barley HVA22. Meanwhile, most PeHVA22s exhibited induced expression post-drought treatment but were suppressed under salt, low and high-temperature conditions, indicating a unique role in drought response. Additionally, PeHVA22s displayed tissue-specific expression patterns across diverse tissues, except for PeHVA22B which maybe a pseudogene. Notably, PeHVA22C, PeHVA22E, and PeHVA22F predominantly expressed in fruit, indicating their involvement in fruit development. Almost all PeHVA22s showed variable expression at different developmental stages of stamens or ovules, implying their roles in passion fruit's sexual reproduction. The intricate roles of PeHVA22s may result from diverse regulatory factors including transcription factors and CREs related to plant growth and development, hormone and stress responsiveness. These observations highlighted that PeHVA22s might play conserved roles in ABA response and drought stress tolerance, and also be participated in the regulation of passion fruit growth and floral development.

[Construction of prokaryotic expression vector of HPV16E6 gene and its expression].

Prokaryotic expression vector of mouse HPV16E6 gene was constructed. A pair of primers were designed according to the digestion sites in plasmid pGEX-KG and the HPV16E6 gene sequence published by GenBank. The DNA fragment of 321bp was amplified by PCR from the HPV recombinant plasmid with HPV16E6 gene, then cloned into pGEX-KG and transformed into the host E. coli strain JM109. The fragment was conformed to the original sequence, which indicated that fusion expression vector pGEX-KG-HPV16E6 was constructed. The pGEX-KG-HPV16E6 plasmid was taken and transformed into BL21(DE3) for expression. Induced by IPTG at 37 degrees C, the expression product of HPV16E6 gene was identified by SDS-PAGE and Western blot. HPV16E6 fusion protein had been expressed successfully in the form of inclusion bodies, the molecular weight of fusion protein being 38 kD. Meanwhile, the optimum condition of HPV16E6 fusion protein expression was induced with 1.0 mmol/L IPTG for 4h. The fusion protein reacted specifically with the antibodies against HPV16E6. HPV16E6 gene was successfully expressed in E. coli, which could be used as a basis for preparing HPV16E6 vaccine in human.

[Prokaryotic expression of HPV 16L1 and optimization of expression conditions].

OBJECTIVE: To construct the HPV16 L1 prokaryotic expression plasmid and to optimize its expression. METHODS: A pair of primers was designed according to plasmid sequences of pGEX-KG and the HPV16L1 genes published by GeneBank. The DNA fragment of 1500 bp was amplified by PCR from the HPV recombinant plasmid with HPV16L1 gene, then cloned into pGEX-KG and transformed into the host E.coli strain JM109. The pGEX-KG-HPV16L1 plasmid was taken and transformed into BL21(DE3) for expression. Induced by IPTG at 37 degree, the expression product of HPV16L1 gene was identified by SDS-PAGE and Western blot. RESULTS: HPV16L1 fusion protein was expressed successfully in the form of inclusion bodies. The molecular weight was 83 kD. Meanwhile, the optimum condition of HPV16L1 fusion protein expression was induced with 1.0 mmol*L(-1) IPTG for 4 h. The fusion protein reacted specifically with antibodies against HPV16L1. CONCLUSION: The prokaryotic expression vector of HPV16L1 gene has been constructed and expressed in E.coli successfully.