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浙江大学学报（农业与生命科学版）
浙江大学学报（农业与生命科学版）  2020, Vol. 46 Issue (4): 407-416    DOI: 10.3785/j.issn.1008-9209.2019.09.291
园艺学     
紫玉兰MlSOC1基因亚细胞定位及花芽分化时期的表达分析
宣铃娟(),程少禹,戴梦怡,王卓为,申亚梅()
浙江农林大学风景园林学院，杭州 311300
Subcellular localization and expression analysis of MlSOC1 genes during flower bud differentiation period in Magnolia liliflora
Lingjuan XUAN(),Shaoyu CHENG,Mengyi DAI,Zhuowei WANG,Yamei SHEN()
College of Landscape Architecture, Zhejiang A & F University, Hangzhou 311300, China
 全文: PDF(8795 KB)   HTML
摘要：

为了探究SOC1基因在紫玉兰（Magnolia liliflora）成花过程中的作用，通过对景宁木兰转录组数据（NCBI数据库编号：SRP129819）进行筛选，利用同源克隆的方法得到2个SOC1基因的编码区序列；运用在线生物信息学分析工具对这2个基因进行序列分析，并结合实时荧光定量聚合酶链式反应（real-time fluorescence quantitative polymerase chain reaction, qRT-PCR）分析这2个基因在紫玉兰不同花芽分化期的表达模式。将获得的2个SOC1基因分别命名为MlSOC1-1和MlSOC1-2，其中：MlSOC1-1长度为666 bp，编码221个氨基酸；MlSOC1-2长度为654 bp，编码217个氨基酸。2个SOC1基因都具有保守的SOC1基序，属于SOC1/TM3亚家族基因。系统进化树分析结果显示，MlSOC1-1与皱叶木兰（Magnolia praecocissima）中的SOC1同源基因亲缘关系最近，而MlSOC1-2与北美木兰（M. virginiana）中的SOC1同源基因亲缘关系最近，2个基因都与木兰科木兰属植物的遗传距离最近。亚细胞定位试验结果发现，MlSOC1-1和MlSOC1-2都被定位于细胞核上。表达分析结果表明：MlSOC1-1与MlSOC1-2基因相比，前者除了参与紫玉兰芽的成花转变外，还有可能对紫玉兰花器官合成有一定作用。通过对紫玉兰2个MlSOC1基因的研究，发现2个MlSOC1基因在花芽分化过程中的作用存在差异，这为进一步研究MlSOC1基因在紫玉兰成花过程中的作用提供了理论基础。
关键词： 紫玉兰;  MlSOC1基因;  花芽分化;  亚细胞定位;  表达分析    
Abstract:

In order to clarify the mechanism of SOC1 inthe process of flower formation in Magnolia liliflora, two SOC1 genes were screened out based on the transcriptome datain M. sinostellata, and the complete coding regions of these two SOC1 genes were obtained by homologous gene cloning. The bioinformatics software was used to analyze the information of these two genes, and the expression patterns in different flower bud differentiation periods were verified by real-time fluorescence quantitative polymerase chain reaction (qRT-PCR). The results showed that the length of MlSOC1-1 was 666 bp, which encoding 221 amino acids, and the length of MlSOC1-2 was 654 bp, which encoding 217 amino acids. Both of these two MlSOC1 genescontained highly conservative SOC1 motifs, which indicated that the genes belong to the SOC1/TM3 subfamily. Phylogenetic tree analysis showed that MlSOC1-1 was closely related to SOC1 homologous gene of M. praecocissima, while MlSOC1-2 was closely related to SOC1 homologous gene of M. virginiana. Subcellular localization experiments revealed that both genes were located on the nucleus. Besides, the results of expression analysis showed that compared with MlSOC1-2, in addition to participating in the flower transformation of the flower bud in M. liliflora, MlSOC1-1 might also play a role in the flower opening and the synthesis of flower organs. These results show that there are differences in the roles of these two MlSOC1 genes in flower bud differentiation, which provides a theoretical basis for further study of the role of MlSOC1 genes in flower formation of M. liliflora.
Key words: Magnolia liliflora    MlSOC1 gene    flower bud differentiation    subcellular localization    expression analysis
收稿日期: 2019-09-29 出版日期: 2020-09-11
CLC:  Q 785  
基金资助: 浙江省科技计划项目(2019C02023)
通讯作者: 申亚梅     E-mail: 1075855127@qq.com;yameishen@zafu.edu.cn
作者简介: 宣铃娟（https://orcid.org/0000-0003-4196-6402），E-mail：1075855127@qq.com
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引用本文:

宣铃娟,程少禹,戴梦怡,王卓为,申亚梅. 紫玉兰MlSOC1基因亚细胞定位及花芽分化时期的表达分析[J]. 浙江大学学报（农业与生命科学版）, 2020, 46(4): 407-416.

Lingjuan XUAN,Shaoyu CHENG,Mengyi DAI,Zhuowei WANG,Yamei SHEN. Subcellular localization and expression analysis of MlSOC1 genes during flower bud differentiation period in Magnolia liliflora. Journal of Zhejiang University (Agriculture and Life Sciences), 2020, 46(4): 407-416.

链接本文:

http://www.zjujournals.com/agr/CN/10.3785/j.issn.1008-9209.2019.09.291    或    http://www.zjujournals.com/agr/CN/Y2020/V46/I4/407

MlSOC1基因功能

Function of MlSOC1 gene

引物名称

Primer name

引物序列（5′→3′）

Primer sequence (5′→3′)

基因克隆

Gene cloning of MlSOC1

GcSOC1-1-FATCGCAGCATCTGTAAC
GcSOC1-1-RTTAGGCAGCGAGGTCGA
GcSOC1-2-FCCATTCTCGATCTTGCTG
GcSOC1-2-RCAATGGGTAGCGTGGTA

表达分析

Expression analysis of MlSOC1

qRTSOC1-1-FCTGAGGTGGCATTGATC
qRTSOC1-1-RTCGAACTTCCATTGCTG
qRTSOC1-2-FTAGAGGAACTGCAACAC
qRTSOC1-2-RAACGGTGTTTTCTTCTGATAGG
qRTActin-FACGAATCCGGTCCATCCATT
qRTActin-RCCGTTCCACCAGGCAATATG

载体构建

Vector construction of MlSOC1

VcSOC1-1-FCATGCCATGGATGGTGAGGGGAAAAACGCAGATGA
VcSOC1-1-RGGTACTAGTCCGAGCTTTCATGGGATAACGTTTTTTC
VcSOC1-2-FCATGCCATGGATGGTGAGAGGGAAGACGCAGATGA
VcSOC1-2-RGGTACTAGTCCGACCTTGCAATGGGTAGCGTGTTA
表1  紫玉兰MlSOC1基因克隆、表达分析及载体构建相关引物
图1  紫玉兰花芽分化过程A～F.花芽分化时芽外部形态特征；G～L.花芽分化时芽内部形态特征。Le：叶；Fl：花原基；Sc：鳞毛；Se：萼片原基；Pe：花瓣原基；St：雄蕊原基；Te：花被片；Pi：雌蕊原基。
图2  MlSOC1-1和MlSOC1-2基因核苷酸序列比对
图3  MlSOC1-1和MlSOC1-2氨基酸序列与其他物种SOC1氨基酸序列的比对ACV88635.1：北美木兰；XP_023900830.1：栓皮栎；NP_001267909.1：葡萄；AHI_85950.1：山核桃；NP_001236377.1：大豆。
图4  紫玉兰MlSOC1-1和MlSOC1-2蛋白与其他物种SOC1蛋白的进化树
图5  载体构建示意图1：pCMBIA1302-MlSOC1-1/2载体；2：pCMBIA1302原始载体。
图6  MlSOC1-1/2蛋白亚细胞定位A～C.对照组GFP蛋白亚细胞定位；D～F. MlSOC1-1蛋白亚细胞定位；G～I. MlSOC1-2蛋白亚细胞定位。
图7  MlSOC1-1和MlSOC1-2在不同花芽分化期叶和花芽中的表达分析A～B.叶；C～D.花芽。S1：未分化期；S2：分化初期；S3：萼片原基分化期；S4：花瓣原基分化期；S5：雄蕊原基分化期；S6：雌蕊原基分化期。*表示在P＜0.05水平差异有统计学意义；**表示在P＜0.01水平差异有高度统计学意义；***表示在P＜0.001水平差异有极高度统计学意义。
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