Abstract:
【Objective】
DlSWC5,a gene related to the ATP-dependent chromatin remodeling in longan(
Dimocarpus longan Lour.),was cloned,and its subcellular localization and expression characterization were analyzed,to provide theoretical basis for investigating the regulatory function of this gene during the growth and development,and early stage of somatic embryogenesis process in longan.【Method】Embryogenic callus(EC)of the early stage of somatic embryogenesis(SE)of longan variety Honghezi was used as primary experimental materials in this study,in combination with three generations of longan genome data,cloned the cDNA sequence of longan
DlSWC5 gene by RT-PCR,conducted the bioin‐formatics analysis,predicted the interaction regulation network of DlSWC5 protein based on STRING database,and verified protein subcellular localization by laser scanning confocal experiment. Transcriptome sequencing and real-time fluorescence quantitative PCR were respectively used to detect the expression of
DlSWC5 gene in the early stage of somatic embryogenesis and different tissues,and in EC under different auxin(IAA)concentration treatments.【Result】The cDNA sequence of DlSWC5 gene was cloned,which existing 99.80% similarity with CDS sequence of DlSWC5 gene in longan genome,and encoded 340 amino acid residues. DlSWC5 protein,without signal peptide and transmembrane structure, contained a total of 44 phosphorylation sites,its molecular formula was C
1622H
2599N
459O
542S
8,its relative molecular weight was 37458.71 Da,its isoelectric point (pI)was 5.74. The predicted result of DlSWC5 protein structure showed that its secondary structure contained 48.53% α-helix,40.29% random coil,6.47% extended strand and 4.71% β-turn,and its tertiary structure was highly consistent with the homologous protein of
Acer yangbiense. The prediction of protein interaction showed that DlSWC5 protein could interact with 10 proteins such as PIE1,SWC2 and SWC4. Subcellular localization showed that DlSWC5 protein was located in the nucleus. There was no obvious difference in the relative expression level of
DlSWC5 gene during the early stage of somatic embryogenesis in longan. The expression of
DlSWC5 gene was detected in the early stage of somatic embrgenesis and seeds,roots,stems,leaves,buds,flowers,pericarp,pulp and young fruits. Compared with the control group(0 mg/L),the relative expression level of
DlSWC5 gene was significantly decreased under 0.5,1.0,1.5 and 2.0 mg/L IAA treatments for 24 h(
P<0.05).【Conclusion】
DlSWC5 gene has been successfully cloned,and its encoded protein is unstable hydrophilic protein,which is acidic and localized in the nucleus. The expression of
DlSWC5 gene is down-regulated by IAA induction in EC stage of the early stage of somatic embrgenesis.