Screening of EMS-Induced Drought-Tolerant Sugarcane Mutants Employing Physiological, Molecular and Enzymatic Approaches
Abstract
:1. Introduction
2. Materials and Methods
2.1. Plant Source and In Vitro Callus Regeneration
2.2. Measurement of Optimal EMS Concentration and Mutagenic Treatment
2.3. Measurement of Optimal In Vitro PEG-6000.Stimulated Osmotic Stress
2.4. Physiological Index Detection
2.4.1. Chlorophyll a
2.4.2. Leaf Chlorophyll Content
2.4.3. Photosynthetic Rate
2.5. DNA Isolation
2.6. SSR Marker PCR Amplification and CELL-1 Enzyme Digestion
3. Results
3.1. Effect of Various Concentrations of EMS on Sugarcane Calli Differentiation
3.2. Effect of Various Concentrations of PEG-6000-Stimulated Stress on Seedling Growth
3.3. Physiological Analysis
3.4. SSR Analysis
4. Discussion
5. Conclusions
Author Contributions
Acknowledgments
Conflicts of Interest
References
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No. | SSR Name | Repeat Motif | Tm (°C) | F-Primer | R-Primer | Result |
---|---|---|---|---|---|---|
1 | SMC336BS | (TG)23(AG)19 | 62 | ATTCTAGTGCCAATCCATCTCA | CATGCCAACTTCCAAACAGAC | 5 |
2 | SMC569CS | (TG)37 | 62 | GCGATGGTTCCTATGCAACTT | TTCGTGGCTGAGATTCACACTA | 0 |
3 | SMC31CUQ | (TC)10(AC)22 | 62 | CATGCCAACTTCCAATACAGACT | AGTGCCAATCCATCTCAGAGA | 5 |
4 | SMC334BS | (TG)36 | 60 | CAATTCTGACCGTGCAAAGAT | CGATGAGCTTGATTGCGAATG | 5 |
5 | Sep-17 | (CCT)9 | 60 | ACCCTGCTGGTCTCCTCC | ACGTTCGACGTCGTGTAGTG | 5 |
6 | Sep-23 | (AGA)9 | 60 | GTGTTCAGGCAGATGGTCCT | GTCGATGGCACCGATTTATT | 5 |
7 | Sep-59 | (CGG)9 | 60 | TTCCATTTACTCCTCCGTGC | CTCCCCCTCCTCGTACTTGT | 5 |
8 | Sep-6 | (GCA)5(ACA)13 | 60 | CAGCCCATTAACCAAGCAAT | GAAGCAGCTGTTGCTCACTG | 5 |
9 | Sep-70 | (GCG)8 | 60 | AACTCACCCAACAAAGCGAC | AGACGAAGAGCTCGTGGATG | |
10 | Sep-8 | (CT)17 | 60 | CTTGCTTCCCCTTTACTCCC | GAGGCGCCTTACTGTTCTTG | 5 |
11 | Sep-84 | (GGT)9 | 60 | AGAGACCGTAATGGTGACCG | ACCACCACCACCACCATACT | 5 |
12 | Sep-89 | (CGT)10 | 60 | AGCTCTGATTTTTGGGGGTT | GGAAGACAGTGGACGAGGTC | 5 |
13 | SMC286CS | (TG)43 | 58 | TCAAATGGGACCTTATTGGAG | TCCCTCGATCTCCGTTGTT | 5 |
14 | SMC119CG | (TTG)12 | 58 | TTCATCTCTAGCCTACCCCAA | AGCAGCCATTTACCCAGGA | 5 |
15 | mSSCIR43 | (GT)3(AT)2(GT)29 | 52 | ATTCAACGATTTTCACGAG | AACCTAGCAATTTACAAGAG | 0 |
ROC22 | FN39 | ||||||
---|---|---|---|---|---|---|---|
Treatment Time (h) | Treatment Concentration (%) | Number of Germinated Calli | Relative Differentiation Rate (%) | Treatment Time (h) | Treatment Concentration (%) | Number of Germinated Calli | Relative Differentiation Rate (%) |
5 | 0 | 84 | 100 | 5 | 0 | 85 | 100 |
0.05 | 80 | 95.2 | 0.05 | 81 | 95.2 | ||
0.1 | 70 | 83.3 | 0.1 | 70 | 84.7 | ||
0.2 | 48 | 57.1 | 0.2 | 49 | 57.6 | ||
0.5 | 30 | 35.7 | 0.5 | 31 | 36.4 | ||
10 | 0 | 83 | 100 | 10 | 0 | 84 | 100 |
0.05 | 78 | 93.9 | 0.05 | 77 | 91.6 | ||
0.1 | 56 | 67.4 | 0.1 | 53 | 63.0 | ||
0.2 | 34 | 40.9 | 0.2 | 28 | 33.3 | ||
0.5 | 9 | 10.8 | 0.5 | 5 | 5.9 | ||
15 | 0 | 83 | 100 | 15 | 0 | 82 | 100 |
0.05 | 76 | 91.5 | 0.05 | 70 | 85.3 | ||
0.1 | 45 | 54.2 | 0.1 | 39 | 47.5 | ||
0.2 | 21 | 25.3 | 0.2 | 4 | 4.8 | ||
0.5 | 4 | 4.8 | 0.5 | 0 | 0 | ||
20 | 0 | 80 | 100 | 20 | 0 | 83 | 100 |
0.05 | 69 | 86.2 | 0.05 | 66 | 79.5 | ||
0.1 | 29 | 36.2 | 0.1 | 24 | 28.9 | ||
0.2 | 3 | 3.7 | 0.2 | 0 | 0 | ||
0.5 | 0 | 0 | 0.5 | 0 | 0 | ||
25 | 0 | 81 | 100 | 25 | 0 | 79 | 100 |
0.05 | 66 | 81.4 | 0.05 | 60 | 75.9 | ||
0.1 | 9 | 11.1 | 0.1 | 6 | 7.5 | ||
0.2 | 0 | 0 | 0.2 | 0 | 0 | ||
0.5 | 0 | 0 | 0.5 | 0 | 0 |
Genotypes | Fv/Fm | Diff b/w Chl Content Before and After Trt | Photosynthetic Rate |
---|---|---|---|
ROC22 | 0.77 | 0.22 | 4.89 |
MR22-1 | 0.75 | 0.15 | 3.88 |
MR22-2 | 0.75 | 0.43 | 7.84 |
MR22-3 | 0.72 | 0.16 | 5.05 |
MR22-4 | 0.76 | −0.08 | 4.55 |
MR22-5 | 0.74 | −0.22 | 2.45 |
MR22-6 | 0.76 | 0.28 | 7.36 |
MR22-7 | 0.78 | 0.27 | 0.95 |
MR22-8 | 0.76 | 0.43 | 4.31 |
MR22-9 | 0.78 | 0.13 | 2.16 |
MR22-10 | 0.76 | −0.07 | 7.13 |
MR22-11 | 0.77 | 0.26 | 0.52 |
MR22-12 | 0.75 | 0.01 | 2.98 |
MR22-13 | 0.76 | 0.61 | 4.05 |
MR22-15 | 0.69 | −0.04 | 10.65 |
MR22-17 | 0.75 | 0.19 | 2.39 |
MR22-20 | 0.73 | −0.14 | 12.13 |
MR22-25 | 0.74 | 0.04 | 7.70 |
FN 39 | 0.77 | 0.39 | 3.38 |
FN39-1 | 0.76 | 0.55 | 3.26 |
FN39-2 | 0.74 | 0.52 | 2.85 |
Genotypes | Average | 5% Sig Level | 1% Sig Level |
---|---|---|---|
ROC 22 | 0.77 | a | a |
MR22-15 | 0.69 | bc | bc |
MR22-20 | 0.73 | c | c |
Genotypes | Average | 5% Sig Level | 1% Sig Level |
---|---|---|---|
ROC22 | 4.89 | a | a |
MR22-2 | 7.8367 | b | a |
MR22-7 | 0.9489 | b | b |
MR22-9 | 2.1615 | b | a |
MR22-11 | 0.5202 | b | b |
MR22-15 | 12.1331 | b | bc |
MR22-20 | 10.6471 | b | bc |
Sample File Name | Size | Height | Sample File Name | Size | Height |
---|---|---|---|---|---|
Sep-23-15P | 141.38 | 1542 | Sep-23-20P | 145.41 | 4215 |
Sep-23-15P | 168.83 | 841 | Sep-23-20P | 150.9 | 466 |
Sep-23-15P | 174.83 | 123 | Sep-23-20P | 163.05 | 465 |
Sep-23-15P | 182.96 | 142 | Sep-23-20N | 146.48 | 30783 |
Sep-23-15P | 201.94 | 150 | Sep-23-20N | 150.9 | 9531 |
Sep-23-15P | 208.33 | 119 | Sep-23-20N | 163.06 | 6201 |
Sep-23-15N | 142.03 | 210 | |||
Sep-23-15N | 146.62 | 92 | |||
Sep-23-15N | 159.06 | 77 |
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Khalil, F.; Naiyan, X.; Tayyab, M.; Pinghua, C. Screening of EMS-Induced Drought-Tolerant Sugarcane Mutants Employing Physiological, Molecular and Enzymatic Approaches. Agronomy 2018, 8, 226. https://doi.org/10.3390/agronomy8100226
Khalil F, Naiyan X, Tayyab M, Pinghua C. Screening of EMS-Induced Drought-Tolerant Sugarcane Mutants Employing Physiological, Molecular and Enzymatic Approaches. Agronomy. 2018; 8(10):226. https://doi.org/10.3390/agronomy8100226
Chicago/Turabian StyleKhalil, Farghama, Xiao Naiyan, Muhammad Tayyab, and Chen Pinghua. 2018. "Screening of EMS-Induced Drought-Tolerant Sugarcane Mutants Employing Physiological, Molecular and Enzymatic Approaches" Agronomy 8, no. 10: 226. https://doi.org/10.3390/agronomy8100226