Subcellular Localization and Coding Gene Expression Pattern Analysis of NtPsbS in Tobacco

doi:10.7668/hbnxb.20195402

Abstract

Abstract:

In order to reveal the expression patterns of PsbS protein in Nicotiana tabacum,the full-length NtPsbS gene sequence was cloned from cDNA of the tobacco cultivar K326,and the multiple protein sequences of NtPsbS gene and PsbS genes of seven crops including rice,tomato and soybean were compared by DNAMAN software.The phylogenetic analysis was carried out by MEGA 11 software,and the phylogenetic tree was established by neighbor-joining method.Tissue expression of NtPsbS gene in tobacco at different growth stages was detected by qRT-PCR.The plant expression vector pS1300-PsbS-GFP was constructed to find subcellular localization of the NtPsbS mature protein.Finally,the mRNA levels of NtPsbS under abiotic stresses were analyzed in K326.The results showed that the total length of tobacco NtPsbS gene was 825 bp,encoding 274 amino acids.The homology of tobacco NtPsbS protein and tomato SlPsbS protein was the highest up to 91%.The NtPsbS gene exhibited the highest mRNA levels in the leaves among the tissues,including leaves,roots,stems,seeds and other parts of K326.The mature NtPsbS protein was located in chloroplasts.For the abiotic stress,it was found that the expression level of NtPsbS was significantly increased under salt stress,cold stress and Abscisic acid(ABA)treatments.In summary,the expression level of NtPsbS was the highest in tobacco leaves at different growth stages,and it was higher when under the treatments of salt stress,cold stress and ABA,indicating that this gene may be involved in the salt and cold stress resistance and ABA metabolic pathway in tobacco,which could provide basis and reference for the functional analysis of the NtPsbS gene in future.

Key words: Nicotiana tabacum, PsbS gene, Subcellular localization, Tissue expression pattern, Abiotic stress

FAN Puqing, ZHOU Houliang, SONG Shanshan, LIN Faming, SHI Yongchun, WANG Xiaoran, WANG Ran, ZHANG Xiaoquan. Subcellular Localization and Coding Gene Expression Pattern Analysis of NtPsbS in Tobacco[J]. Acta Agriculturae Boreali-Sinica, 2025, 40(2): 49-55. doi: 10.7668/hbnxb.20195402.

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Figures/Tables 6

Tab.1 Primer sequences and uses

引物名称 Primer name	引物序列 (5'—3') Primer sequence (5'—3')	用途 Use
NtPsbS	F;ATGGCTCAAACAATGTTGCTGAC	基因克隆
	R:TCACTCCTCTTCCTCATCAG
qRT-NtPsbS	F:CTCTTCCTTTGAAATACCCTTC	检测NtPsbS表达水平
	R:TGAAACCAATTCCTCCTGAGG
qRT-NtL25	F:ATGGCTCAAACAATGTTGCTGAC	检测NtL25表达水平
	R:ATGGCTCAAACAATGTTGCTGAC
NtPsbS-GFP	F:CAGGGGCCCGGGGTCGACATGGCTCAAACAATGTTGCTGAC	亚细胞定位
	R:CCCTTGCTCACCATGGTACCTCACTCCTCTTCCTCATCAG

Fig.1 Comparison of NtPsbS protein and homologous protein sequences (A) and construction of phylogenetic tree(B) AtPsbS.NP_001319163.1;GhPsbS.XP_016708802.1;GmPsbS.XP_003523444.1;NtPsbS.NP_001312190.1;OsPsbS.XP_01521169.1;S1PsbS.NP_001296186.1;TaPsbS.XP_020167324.1;ZmPsbS.NP_001105228.3.

Fig.2 Electrophoresis map of NtPsbS PCR product (A),schematic diagram of pS1300-PsbS-GFP expression vector (B)and expression vector enzyme digestion identification (C)

Fig.3 Expression profile of NtPsbS gene in tissues Different lowercase letters on the column indicate significant differences (P<0.05).The same as Fig.5.

Fig.4 Subcellular localization of NtPsbS-GFP in leaves of Nicotiana benthamiana

Fig.5 Expression level of NtPsbS gene under abiotic stresses

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