Chinese Bulletin of Botany ›› 2020, Vol. 55 ›› Issue (2): 137-146.DOI: 10.11983/CBB19109
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Yang Zhang,Huajie Liu,Ruili Xue,Haixia Li,Hua Li()
Received:
2019-06-14
Accepted:
2019-11-05
Online:
2020-03-01
Published:
2020-02-12
Contact:
Hua Li
Yang Zhang,Huajie Liu,Ruili Xue,Haixia Li,Hua Li. Cloning of Wheat TaLCD Gene and Its Regulation on Osmotic Stress[J]. Chinese Bulletin of Botany, 2020, 55(2): 137-146.
Gene name | Primer sequences | |
---|---|---|
Forward (5'-3') | Reverse (5'-3') | |
TaDCD | GAGGAAGGACGGAAGCCATAT | TCAGGGTCATCGCAAACAGAG |
TaLCD | TCCATTACGCCTACGGAGCAG | CAAGCCGGACCTTACGACCA |
ABF2 | ATCAGAAGGGATAGGGAAGAGTAAT | TTGGTCTGCCGTGAATATCTGT |
HAB1 | GTGTTCTCGCCATGTCTAGGTC | CTATTTCGCAGACTTCTTGGTTG |
HAB2 | GCAGAAGTCCTTATTGCGAGTC | CTCAGAAGTTGCCACCTCCATA |
ABI1 | TGACGGCTGTGAAGAGAGTA | CCATCTCACACGCTTCTTCA |
ABI2 | ATTCAGACCATTCACTGACCCTC | GCTCCGTCGCCAGAACAAG |
NCED3 | TGGCTTCTTTCACGGCAAC | ACAACAATGGCGGGAGAGTTT |
COR47 | GAGGTTACGGATCGTGGAT | GAGCTGTTGGATCGGTGA |
RAB18 | ATGGCGTCTTACCAGAACCGTCCA | TACCCTTGGCCACCTGATCC |
RD29A | GTTACTGATCCCACCAAAGAAGA | GGAGACTCATCAGTCACTTCCA |
RD22 | AGGGCTGTTTCCACTGAGG | CACCACAGATTTATCGTCAGACA |
Actin2 | TACCCGATGGGCAAGTCA | TGCTCATACGGTCAGCGATA |
Table 1 Primers used for qRT-PCR analysis
Gene name | Primer sequences | |
---|---|---|
Forward (5'-3') | Reverse (5'-3') | |
TaDCD | GAGGAAGGACGGAAGCCATAT | TCAGGGTCATCGCAAACAGAG |
TaLCD | TCCATTACGCCTACGGAGCAG | CAAGCCGGACCTTACGACCA |
ABF2 | ATCAGAAGGGATAGGGAAGAGTAAT | TTGGTCTGCCGTGAATATCTGT |
HAB1 | GTGTTCTCGCCATGTCTAGGTC | CTATTTCGCAGACTTCTTGGTTG |
HAB2 | GCAGAAGTCCTTATTGCGAGTC | CTCAGAAGTTGCCACCTCCATA |
ABI1 | TGACGGCTGTGAAGAGAGTA | CCATCTCACACGCTTCTTCA |
ABI2 | ATTCAGACCATTCACTGACCCTC | GCTCCGTCGCCAGAACAAG |
NCED3 | TGGCTTCTTTCACGGCAAC | ACAACAATGGCGGGAGAGTTT |
COR47 | GAGGTTACGGATCGTGGAT | GAGCTGTTGGATCGGTGA |
RAB18 | ATGGCGTCTTACCAGAACCGTCCA | TACCCTTGGCCACCTGATCC |
RD29A | GTTACTGATCCCACCAAAGAAGA | GGAGACTCATCAGTCACTTCCA |
RD22 | AGGGCTGTTTCCACTGAGG | CACCACAGATTTATCGTCAGACA |
Actin2 | TACCCGATGGGCAAGTCA | TGCTCATACGGTCAGCGATA |
Figure 1 Sequence analysis of TaLCD (A) Comparison of the derived amino acid sequences of TaLCD in Triticum aestivum with other 4 species (Aegilops tauschii, Brachypodium distachyon, Setaria italica, and Oryza brachyantha); (B) Phylogenetic analysis of LCD in five species; (C) Schematic structures of TaLCD, black box indicates Aminotran_5 domain.
Figure 2 The expression of TaLCD and enzyme activity of TaLCD in wheat under NaCl and PEG treatments (A) The expression of TaLCD under NaCl treatment; (B) The expression of TaLCD under PEG treatment; (C) The enzyme activity of TaLCD under NaCl treatment; (D) The enzyme activity of TaLCD under PEG treatment. * indicate significant differences (P<0.05).
Figure 3 Expression of TaLCD and enzyme activity of TaLCD in transgenic Arabidopsis (A) RT-PCR; (B) qRT-PCR; (C) Enzyme activity of TaLCD under NaCl treatment. Marker: DNA marker; WT: Wild type; 2-8, 3-4, 5-5, and 12-4: TaLCD transgenic lines; * indicate significant differences (P<0.05).
Figure 4 Seed germination rate of Arabidopsis lines expressing TaLCD under different concentrations of NaCl treatment WT: Wild type; OX-LCD-1 and OX-LCD-2: Transgenic lines; * indicate significant differences (P<0.05).
Figure 5 Root growth of Arabidopsis lines expressing TaLCD under different concentrations of mannitol treatment (A) Phenotype of root growth (Bars=1 cm); (B) Root elongation measurements. WT: Wild type; OX-LCD-1 and OX-LCD-2: Transgenic lines; * indicate significant differences (P<0.05).
Figure 6 Drought resistance of Arabidopsis lines expressing TaLCD (A) Phenotype of three week-old wild type (WT), OX-LCD-1 and OX-LCD-2 plants after drought treatment (Bars=2 cm); (B) Leaf water loss
Figure 7 Seed germination and growth of Arabidopsis lines expressing TaLCD under different concentrations of ABA treatment (A) Phenotype on the 10th day (Bars=1 cm); (B) Seed germination rate after 4 d incubation; (C) Phenotype of root growth on the 10th day (Bars=0.4 cm); (D) Root length on the 10th day. WT: Wild type; ABA: Abscisic acid; * indicate significant differences (P<0.05).
Figure 8 The expression of stress response genes and ABA signaling related genes in Arabidopsis lines expressing TaLCD under drought stress (A)-(D) Stress response genes; (E)-(J) ABA signaling related genes. WT: Wild type; OX-LCD-2: Transgenic lines
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