Identification,Expression and Bioinformatics Analysis of Smoothened Gene of Apis cerana cerana

doi:10.7668/hbnxb.20194415

Abstract

Abstract:

The aim of this study was to clarify the sequence characteristics and expression properties of the Smoothened gene in Apis cerana cerana.The DNA sequence of AcerSmo was amplified and identified,its structure and function were analysed using various bioinformatics software,and the expression characteristics were examined by RT-qPCR for different tissues and developmental periods of Apis cerana cerana.Sequence analysis showed that the AcerSmo was 3 495 bp(GenBank accession number:OP616035),the open reading frame(ORF)region was 2 952 bp,encoding 936 amino acids.The molecular weight of this protein was 111.15 ku and the predicted isoelectric point was 8.82,which had a signal peptide containing 21 amino acids at the N-terminus,and there were 6 transmembrane structures domains.Subcellular localization mostly takes place in the nucleus and endoplasmic reticulum.The secondary and tertiary structures of this protein were dominated by irregular coiled(50.76%),followed by α-helix(25.33%).The sequence of AcerSmo protein sequence was highly conserved from insects to mammals.The results of RT-qPCR showed that AcerSmo was expressed in different tissues of Apis cerana cerana at the age of 30 days and the expression level of AcerSmo in the antennae and head were significantly higher than in other tissues,and in the tissues of the antennae and head of the Apis cerana cerana,the mRNA expression of AcerSmo was the highest at 5 days of age.AcerSmo has the conserved structure of Smoothened protein and was highly expressed in high abundance in antennae,which may play an important role in the olfactory recognition process of honey bees.

Key words: Apis cerana cerana, Smoothened, Sequence characterization, Bioinformatics analysis, Expression characteristics

YU Diandian, SHEN Hongying, WANG Jue, GUO Yuan, GUO Lina. Identification,Expression and Bioinformatics Analysis of Smoothened Gene of Apis cerana cerana[J]. Acta Agriculturae Boreali-Sinica, 2024, 39(3): 230-238. doi: 10.7668/hbnxb.20194415.

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Figures/Tables 11

Tab.1 Primer sequence formation

引物名称 Primer name	引物序列(5'—3') Primer sequence(5'—3')	用途 Purpose
P1	F: TAATATTTGTTCATTATTTCTAAAATTTCC	序列扩增
	R: AGCCCAAGCAATAAAAGAATGAAT
P2	F: TAATATTTGTTCATTATTTCTAAAATTTCC
	R: AGCCCAAGCAATAAAAGAATGAAT
P3	F: ATTTCTAGCAAATGCTTAAAGCCT
	R: GTAAAGCAGCTGCCCATGTTG
P4	F: CATTCGCAAAACGGAAAACATT
	R:GAAAAATTAGTGTTACGTAGTAATATTTTTG
qAcerSmo	F: TGGCAGCTATGGTGTGGTTT	荧光定量
	R: GACAGTGAGGACAAGTGGCA
qArp1	F: TGCTGCACTCGTAGTTGACAATGG
	R: ACCCTGGTGGCGTGGTCTTC

Fig.1 Electrophoresis of the PCR products of AcerSmo M.3000 Marker;1—3.PCR product.

Fig.2 AcerSmo ORF region and deduced amino acid sequence

Fig.3 Predicted signal peptide and transmembrane of AcerSmo A.Predicted signal peptide;B.Transmembrane prediction.

Fig.4 The secondary and tertiary structure prediction of AcerSmo A.Secondary structure prediction;B.The tertiary structure prediction;C.Ramachandran map analysis of the tertiary structural model.

Fig.5 Motif and conserved domain prediction of AcerSmo A.Motif prediction;B.The prediction of conserved domain.

Fig.6 Multiple alignment of AcerSmo

Fig.7 Phylogenetic tree Data in the upper branches are bootstrap of>50% from 1 000 repeats.

Fig.8 Relative expression of AcerSmo in different tissues of 30-day-old Apis cerana cerana AN.Antennae;HE.Head(without antenna);T.Thorax;AB.Abodem;L.Legs;W.Wing.Different lowercase letters indicate significant differences in different tissues(P<0.05).

Fig.9 Relative expression of AcerSmo in the antennae of Apis cerana cerana at different days Different lowercase letters indicated significant differences in different days(P<0.05).The same as Fig.10.

Fig.10 Relative expression of AcerSmo in the antennae of Apis cerana cerana at different days

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