Scientia Agricultura Sinica ›› 2019, Vol. 52 ›› Issue (22): 3976-3986.doi: 10.3864/j.issn.0578-1752.2019.22.004

• MOLECULAR GENETICS • Previous Articles     Next Articles

Response of Receptor-Like Protein Kinase Gene SiRLK35 of Foxtail Millet to Salt in Heterologous Transgenic Rice

LI XiaoBo1,2,LI Zhen2,DAI ShaoJun3,PAN JiaoWen2,WANG QingGuo2,GUAN YanAn4,5,DING GuoHua1(),LIU Wei2,5()   

  1. 1 School of Life Sciences and Technology, Harbin Normal University, Harbin 150025
    2 Biotechnology Research Center, Shandong Academy of Agricultural Sciences/Shandong Provincial Key Laboratory of Crop Genetic Improvement, Ecology and Physiology, Jinan 250100
    3 College of Life and Environmental Sciences, Shanghai Normal University/Development Center of Plant Germplasm Resources, Shanghai 200234
    4 Crop Research Institute, Shandong Academy of Agricultural Sciences/Shandong Engineering Laboratory for Featured Crops, Jinan 250100
    5 College of Life Sciences, Shandong Normal University, Jinan 250014
  • Received:2019-07-02 Accepted:2019-09-19 Online:2019-11-16 Published:2019-11-16
  • Contact: GuoHua DING,Wei LIU E-mail:hsddgh@hrbnu.edu.cn;wheiliu@163.com

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Abstract:

【Objective】Salt stress affects the crop yield and quality, while the crop salt tolerance is regulated by specific genes. Using heterogeneous over-expressed rice lines (OE-1, OE-2, and OE-3) of foxtail millet receptor-like protein kinase gene SiRLK35, the possible mechanisms of SiRLK35 under salinity will be dissected on phenotypic, physiological and molecular levels. 【Method】 The expressions of SiRLK35 in OE-1, OE-2, and OE-3 were analyzed by qRT-PCR. The phenotypes of three-leaf stage seedlings treated with 0, 150 and 200 mmol·L -1 NaCl were observed, and the lengths of seedlings and roots were measured after treated with 150 mmol·L -1 NaCl for 3 days. The dry weights, death rate and dead leaf rate of four-leaf stage seedlings that treated with 150 mmol·L -1 NaCl for 14 days were also measured. OE-1with the highest SiRLK35 expression was further used for DAB and NBT dyeing analysis. The activities of partial antioxidases, and expression patterns of marker genes were detected. 【Result】 There was the highest SiRLK35 expression level in OE-1. The growth of control and OE seedlings were all inhibited under 150 mmol·L -1 NaCl. The decreased levels of dry weight, the death rate and dead leaf rate of OE rice were all lower than those of the control, together with the less accumulation of O 2- and H2O2, and the higher activities of antioxidases under salinity. Partial of salt responsive genes were up-regulated in SiRLK35 OE lines, especially the OsLEA3 was induced about 1.9 times higher after treated with NaCl for 24 h. 【Conclusion】 SiRLK35 OE rice lines have tolerance under salinity, and the gene SiRLK35 of foxtail millet could participate in salt response by regulation the activities of antioxidases and related signal pathways.

Key words: foxtail millet (Setaria italica L.), SiRLK35, salt tolerance, antioxidase, salt responsive genes

Table 1

Genes information and specific primers"

基因名称Gene name 登录号Accession number 正向引物Forward primer (5'-3') 反向引物Reverse primer (5'-3')
OsACTIN MS971743.1 TCGGCTACAACCCTGACAA CAAACTTGACGGCAATGTGG
SiRLK35 XM_004956247.2 AAATAGTTGGGTAGACGAGG TACGCAGATGATTGACAGAT
OsP5CS D49714.1 CTATTCCTCGTAATGTTGG CTTAGTTGACGCCTTGAT
OsLEA3 Z68090.1 ACAAGGACACCTCTGCCACC AATAGACCCAAAGGGAAATCA
OsNHX1 AB021878.1 GAGAGGAGCTGTGTCGATTG CTCTCGAGGTCAGAACCTTG
OsHKT1 KU501218.1 ACCATAAGCACAACCCAG AATCCTAAGAACCACCTCA
OsNAC6 AB028185.1 GATCATGCACGAGTACCGC GCACCCAATCATCCAACCT
OsDREB2A JQ341059.1 GCTGAGATCCGTGAACCA ACCATACATTGCCCTTGC

Fig. 1

PCR verification of SiRLK35 transgenic rice M: Trans2K Plus DNA Marker; 1: Zhonghua 11; 2-3: OE-1; 4-5: OE-2; 6-7: OE-3"

Fig. 2

The relative expression levels of SiRLK35 in control and transgenic rice lines ***: Indicates that compared with Zhonghua11, the relative expression levels of SiRLK35 in OE rice lines were significantly different at P<0.001 (n = 3). The same as below"

Fig. 3

Salt-resistance detection of control and SiRLK35 transgenic rice lines A: The phenotypes of control and transgenics treated with 0 , 150 and 200 mmol·L-1 NaCl for 2 days, Bar=5 cm; B:Plant height of control and transgenics treated with 0 and 150 mmol·L-1 NaCl for 3days; C: Root length of control and transgenics treated with 0 and 150 mmol·L-1 NaCl for 3days; D: Relative reduction of dry weight in the above-ground part of control and transgenics treated with 0 and 150 mmol·L-1 NaCl for 14 days; E: Relative reduction of dry weight in underground parts of control and transgenics treated with 0 and 150 mmol·L-1 NaCl for 14 days; F: Death rate of control and transgenics treated with 150 mmol·L-1 NaCl for 14 days; G: Dead leaf rate of control and transgenics treated with 150 mmol·L-1 NaCl for 14 days;* significantly different at P< 0.05 level (n = 3); ** significantly different at P< 0.01 level (n = 3). The same as below"

Fig. 4

Detection of reactive oxygen and antioxidant enzyme activities of control and OE-1 after salt treatment A: DAB and NBT staining analysis of control and OE-1 after 3 day of NaCl treatment, Bar=1cm; B: SOD activity analysis of control and OE-1 after salt treatment; C: POD activity of control and OE-1 after salt treatment; the different letters above the curve indicate significant difference (P<0.05) of different treatments(n=3)"

Fig. 5

The relative expressions of salt responsive genes in control and OE-1 under salinity"

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