Selection high sucrose soybean lines from a mutation polulation derive fron ems treatment

THAI NGUYEN UNIVERSITY UNIVERSITY OF AGRICULTURE AND FORESTRY DUONG THI BICH HUONG SELECTION HIGH SUCROSE SOYBEAN LINES FROM A MUTATION POPULATION DERIVE FROM EMS TREATMENT BACHELOR THESIS Study Mode : Full-time Major : Biotechnology Faculty : Biotechnology and Food Technology Batch : 2012 – 2016 Thai Nguyen, 2016/08/20 THAI NGUYEN UNIVERSITY UNIVERSITY OF AGRICULTURE AND FORESTRY DUONG THI BICH HUONG SELECTION HIGH SUCROSE SOYBEAN LINES FROM A MUTATION POPULATION DERIVE FROM EMS TREATMENT BACHELOR THESIS Study Mode : Full-time Major : Biotechnology Faculty : Biotechnology and Food Technology Batch : 2012 – 2016 Supervisors : Prof Lee Jeong-Dong Ph.D Duong Van Cuong _ Ph.D Pham Bang Phuong Thai Nguyen, 2016/08/20 Thai Nguyen University of Agriculture and Forestry Major Biotechnology Student name Duong Thi Bich Huong Student ID DTN1253150047 Thesis Title Selection high sucrose soybean lines from a mutation population derive from EMS treatment Supervisor (s) Prof Lee Jeong Dr Duong Van Cuong Dr Pham Bang Phuong Abstract: Soybean is one of the most important crops Soybean seeds contain about 35% carbohydrate Sucrose content of soybean carbohydrate range from 2.5 – 8.2%, raffinose ranges from 0.1-0.9%, stachyose ranges 1.4-4.1% Sucrose content in soybean seeds is desired to be high because it is a sweetness-imparting component, it helps extends acceptance for soyderived food products To improve of quality carbohydrate, the breeding programs primary focused on increasinge sucrose content Mutagenesis is used to study gene function and obtain new genetic resources for plant breeding to find a new genetic resources We developed EMS –mediated mutant population to select high sucrose concentration lines by megazyme kit to analyze sucrose content Selection of 710 out of 3777 mutant lines derived from a mutation population which was treated EMS 0.3% Finishing the preliminary analysis, as a results we obtained 390 lines with sucrose content of larger than that of Pungsanamul The remaining 320 lines were lower Selected 31 soybean lines have larger than that sucrose content Pungsanamul were selected and reanalyzed Sucrose concentration of all 31 those lines were larger than that Pungsanamul sucrose content PE 1472 was confirmed as a unique germplasm with the highest sucrose content (21.216 g/100g) PE 831(16.818 g/100g) were confirmed with extremely low concentrations Pungsanamul was identified around 16.122g/100g The identified germplasm with high sucrose profiles will be valuable in breeding specialty soybeans for improved sugar content The selection of 31 mutant lines with high sucrose profiles will facilitate in a large-scale soybean breeding program Keywords Number of pages Date of Submission High sucrose, EMS, soybean mutaion 63 2016/08/20 i ACKNLOWLEDGMENTS The firstly, I would like to express the gratitude and profound thanks to special person who always admire and respect That is my wonderful supervision Professor, Jeong-Dong Lee You was so kind, love the students, caring, sharing, listening, and always willing to what's best for students Thank you for having the faith in myself and giving me a gold opportunity to internship during the duration Exchange student’s program in your lab Here I have had many great experiences and learned a lot of experience from the learning environment and the social environment In deep my heart, you are a greatest teacher The secondly, I would also like to give acknowledgements to Park Cheolwoo The day to day operations during my research and life would have been nearly impossible without the guidance Park Cheolwoo, he was by my side every step of the way and provided countless suggestions on how to better my studies I am very thankful for your time, help and support throughout my graduate studies The thirdly, I would to thank my advisor Dr Nguyen Minh Phuc, Master Huyn Jo although different major and university but whenever they always helped, supported, and guided carefully for their guidance and feedback during my academic research They were crucial elements to the success of the entire research team They were always there to bounce ideas off of and give a helping hand wherever they could I have benefited from the work of the groups around Korea over the years and I have enjoyed the friendship and collegiality of the students, staff, and others who collaborated in the group’s projects during this time A big thanks goes out to all my member in my lab Sovetgul Asekova, Bota, Min Su Kim, Jae-eun Jeong, Dong Ho Lee and member in other lab Everyone there was more than willing to help my with my research in any way they could Another great thanks goes out to my parents, Binh and Loan who raised me into the young lady I am today I would not have been able to accomplish so much in my undergraduate studies without their love, support, and guidance, encouragement and motivation ii I would like to start off by thanking God for good health and all the great possibilities that he has put into my life I have been very fortunate in my life and been allowed the opportunity for higher education Lastly, I would like to be special big thank my great supervisor Dr Pham Bang Phuong and Dr Duong Van Cuong at Thai Nguyen University of Agriculture and Forestry who recommended me to be today I have had a big chance to learn and conducting this topic Especially, Dr Pham Bang Phuong always oriented and create the most favorable conditions for my learning path Anytime I have difficulty in writing thesis who gave the best advice to support the shared interest enthusiastically help to anyone Another great thanks goes to all teacher in my faculty: Biotechnology and Food taught knowledge and communication have valuable experience Many thank you and best regards Student Duong Thi Bich Huong iii CONTENTS ACKNLOWLEDGMENTS ii CONTENTS iv LIST OF FIGURES v LIST OF TABLES vi LIST OF ABBREVIATION vii I INTRODUCTION 1.1 Soybean 1.2 The sucrose .4 1.2.1 Structure 1.2.2 The function of sucrose .6 1.2.2.1 The role of sucrose in the body 1.2.2.2 The role of sucrose in food product 1.2.3 Trade and economic .12 1.2.4 Situation research 15 1.2.5 Chemical Mutagenesis 16 1.3 Research Objective .17 II MARERIAL AND METHODS 18 2.1 Material and equipment 18 2.1.1 Plant Material 18 2.1.2 Equipment and chemical 18 2.2 Method .20 2.2.1 Carbohydrates extraction 20 2.2.2 Sucrose quantification by the GOPOD/invertase method (enzymatic method) 21 2.2.3 Calculation 27 III RESULTS AND DISCUSSION 28 3.1 The soybean seeds treated by EMS 28 3.2 Determination carbohydrates extraction 29 3.3 Preliminary evaluation of the sugar content .29 3.4 Selection of high sucrose content lines 34 IV CONCLUSION .41 REFERENCES 42 APPENDICES .46 iv Thai Nguyen University of Agriculture and Forestry Major Biotechnology Student name Duong Thi Bich Huong Student ID DTN1253150047 Thesis Title Selection high sucrose soybean lines from a mutation population derive from EMS treatment Supervisor (s) Prof Lee Jeong Dr Duong Van Cuong Dr Pham Bang Phuong Abstract: Soybean is one of the most important crops Soybean seeds contain about 35% carbohydrate Sucrose content of soybean carbohydrate range from 2.5 – 8.2%, raffinose ranges from 0.1-0.9%, stachyose ranges 1.4-4.1% Sucrose content in soybean seeds is desired to be high because it is a sweetness-imparting component, it helps extends acceptance for soyderived food products To improve of quality carbohydrate, the breeding programs primary focused on increasinge sucrose content Mutagenesis is used to study gene function and obtain new genetic resources for plant breeding to find a new genetic resources We developed EMS –mediated mutant population to select high sucrose concentration lines by megazyme kit to analyze sucrose content Selection of 710 out of 3777 mutant lines derived from a mutation population which was treated EMS 0.3% Finishing the preliminary analysis, as a results we obtained 390 lines with sucrose content of larger than that of Pungsanamul The remaining 320 lines were lower Selected 31 soybean lines have larger than that sucrose content Pungsanamul were selected and reanalyzed Sucrose concentration of all 31 those lines were larger than that Pungsanamul sucrose content PE 1472 was confirmed as a unique germplasm with the highest sucrose content (21.216 g/100g) PE 831(16.818 g/100g) were confirmed with extremely low concentrations Pungsanamul was identified around 16.122g/100g The identified germplasm with high sucrose profiles will be valuable in breeding specialty soybeans for improved sugar content The selection of 31 mutant lines with high sucrose profiles will facilitate in a large-scale soybean breeding program Keywords Number of pages Date of Submission High sucrose, EMS, soybean mutaion 63 2016/08/20 i LIST OF TABLES Table 1: Sucrose concentration both the 48 mutant lines and Pungsanamul were determined by the GOPOD/invertase method .31 Table 2: The first replications, the sucrose concentrations % (g/100 g) The value of glucose content was measured at 510 nm, µg /ml value, µg/mg value were determined by the GOPOD/invetase method in seed of 31 different soybean lines from a mutation population derived from EMS treatment and a control line (Pungsanamul) 35 Table 3: The second replications, the sucrose concentrations % (g/100 g) The value of glucose content was measured at 510 nm,µg /ml value, µg/mg value were determined by the GOPOD/invertase method in seed of 31 different soybean lines from a mutation population derived from EMS treatment and a control line (Pungsanamul) 36 Table 4: Mean two replications of sucrose concentrations in % (g/100 g) were determined by the GOPOD/invetase method In seed of 31 different soybean lines from a mutation population derived from EMS treatment and the control sample Pungsanamul 37 vi LIST OF ABBREVIATION ATP BU DN EMS ELISA FAO FDA g G6DH GO GOPOD HPAE-PAD HPLC mL N NADPH NIRS NSP Rep PCR RMP Pungsan RNA SBP TLC µL USA USD USDA °C °F: % Adenosin triphosphate Bushel Deoxyribonucleic acid Ethyl Methane Sulphonate Enzyme-Linked ImmunoSorbent Assay Food and Agriculture Organization Food and Drug Administration Gram Glucose-6- phosphate dehydrogenase (or G6PDH) Glucose oxidase Glucose oxidase peroxidase High-Performance Anion-Exchange Chromatography-Pulsed Amperometric Detection High pressure liquid chromatography Milliliter Nitrogen Nicotinamide adenine dinucleotide phosphate Near-infrared reflectance spectroscopy Nonstarch polysaccharides Replication Polymerase Chain Reaction Revolutions Per Minute Pungsanamul Ribonucleic acid Sucrose blinding protein Thin layer chromatography Microliter United States Dollar The United States of America United States Department of Agriculture Celsius degree Fahrenheit degree Percent vii I INTRODUCTION 1.1 Soybean Cultivated soybean [G max (L.) Merr] is a diploidized tetraploid (2n=40), in the family Leguminosae, the subfamily Papilionoideae, the tribe Phaseoleae, the genus Glycine Wild and the subgenus Soja (Moench) It is an erect, bushy herbaceous annual that can reach a height of 1.5 meters Three types of growth habit can be found amongst soybean cultivars: determinate, semi-determinate and indeterminate (Bernard and Weiss, 1973) (Figure 1) Figure 1: Soybean plant Cultivation is successful in climates with hot summers, with optimum growing conditions in mean temperatures of 20 to 30 °C (68 to 86 °F); temperatures of below 20 °C and over 40 °C (68 °F, 104 °F) stunt growth significantly They can grow in a wide range of soils, with optimum growth in moist alluvial soils with a good organic Ordinal 111 112 113 114 115 116 117 118 119 120 121 122 123 124 125 126 127 128 129 130 131 132 133 134 135 136 137 138 139 140 141 142 143 144 145 146 147 148 149 150 Entry PE 206 PE 207 PE 208 PE 209 PE 210 PE 211 PE 212 PE 213 PE 215 PE 216 PE 217 PE 218 PE 219 PE 220 PE 221 PE 228 PE 230 PE 232 PE 233 PE 234 PE 236 PE 238* PE 243 PE 245 PE 246 PE 247 PE 248 PE 249 PE 251 PE 252 PE 253 PE 256* PE 258* PE 261 PE 263 PE 265 PE 268 PE 273 PE 280 PE 309 Absorbance at 510 nm 0.756 0.770 0.737 0.772 0.776 0.850 0.825 0.762 0.800 0.827 0.758 0.749 0.734 0.777 0.757 0.961 0.807 1.005 0.886 0.980 0.907 0.957 0.987 0.914 0.907 0.930 0.881 0.968 0.851 0.890 0.857 0.980 1.040 1.018 0.838 0.851 0.720 0.803 0.682 0.899 Con.c (ug/ml) µg/mg 566.923 577.692 552.308 579.231 582.308 639.231 620.000 571.538 600.769 621.538 568.462 561.538 550.000 583.077 567.692 724.615 606.154 758.462 666.923 739.231 683.077 721.538 744.615 688.462 683.077 700.769 663.077 730.000 640.000 670.000 644.615 739.231 785.385 768.462 630.000 640.000 539.231 603.077 510.000 676.923 51.538 52.517 50.210 52.657 52.937 58.112 56.364 51.958 54.615 56.503 51.678 51.049 50.000 53.007 51.608 65.874 55.105 68.951 60.629 67.203 62.098 65.594 67.692 62.587 62.098 63.706 60.280 66.364 58.182 60.909 58.601 67.203 71.399 69.860 57.273 58.182 49.021 54.825 46.364 61.538 49 Percentage of sucrose (g/100g) 9.792 9.978 9.540 10.005 10.058 11.041 10.709 9.872 10.377 10.736 9.819 9.699 9.500 10.071 9.806 12.516 10.470 13.101 11.520 12.769 11.799 12.463 12.862 11.892 11.799 12.104 11.453 12.609 11.055 11.573 11.134 12.769 13.566 13.273 10.882 11.055 9.314 10.417 8.809 11.692 Ordinal Entry 151 152 153 154 155 156 157 158 159 160 161 162 163 165 166 167 168 169 170 171 172 173 174 175 176 177 178 179 180 181 182 183 184 185 186 187 188 189 190 PE 318 PE 320 PE 322 PE 323 PE 324 PE 326 PE 328 PE 329 PE 330 PE 331 PE334 PE 335 PE 342 PE 347 PE 349 PE 354 PE 357 PE 359 PE 361 PE 377 PE 379 PE 383 PE 386 PE 387 PE 404 PE 405 PE 407 PE 411 PE 417 PE 432 PE 437 PE 439 PE 440 PE 441 PE 443 PE 445 PE 446 PE 447 PE 449 Absorbance at 510 nm 0.516 0.688 0.800 0.941 0.970 0.911 0.926 0.892 0.913 0.907 0.896 0.952 0.857 0.662 0.823 0.815 0.792 0.808 0.805 0.793 0.853 0.774 0.832 0.878 0.786 0.741 0.626 0.741 0.665 0.737 0.777 0.738 1.055 0.980 0.958 0.926 0.919 0.967 0.897 Con.c (ug/ml) 382.308 514.615 600.769 709.231 731.538 686.154 697.692 671.538 687.692 683.077 674.615 717.692 644.615 494.615 618.462 612.308 594.615 606.923 604.615 595.385 641.538 580.769 625.385 660.769 590.000 555.385 466.923 555.385 496.923 552.308 583.077 553.077 796.923 739.231 722.308 697.692 692.308 729.231 675.385 50 µg/mg Percentage of sucrose (g/100g) 34.755 6.603 46.783 8.889 54.615 10.377 64.476 12.250 66.503 12.636 62.378 11.852 63.427 12.051 61.049 11.599 62.517 11.878 62.098 11.799 61.329 11.652 65.245 12.397 58.601 11.134 44.965 8.543 56.224 10.683 55.664 10.576 54.056 10.271 55.175 10.483 54.965 10.443 54.126 10.284 58.322 11.081 52.797 10.031 56.853 10.802 60.070 11.413 53.636 10.191 50.490 9.593 42.448 8.065 50.490 9.593 45.175 8.583 50.210 9.540 53.007 10.071 50.280 9.553 72.448 13.765 67.203 12.769 65.664 12.476 63.427 12.051 62.937 11.958 66.294 12.596 61.399 11.666 Oridinal Entry 191 192 193 194 195 196 197 198 199 200 201 202 203 204 205 206 207 208 209 210 211 212 213 214 215 216 217 218 219 220 221 222 223 224 225 226 227 228 229 230 PE 450 PE 457 PE 469 PE 471 PE 472 PE 473 PE 474 PE 475 PE 478 PE 479 PE 480 PE 483 PE 484 PE 488 PE 492 PE 494 PE 500 PE 501 PE 523 PE 524 PE 526 PE 529 PE 535 PE 539 PE 542 PE 544 PE 545 PE 546 PE 549 PE 553 PE 556 PE 560 PE 561 PE 562 PE 567 PE 571 PE 576 PE 588 PE 589 PE 591 Absorbance at 510 nm 1.010 0.921 1.007 0.997 0.801 0.926 0.918 0.978 0.945 0.872 0.860 0.920 0.981 0.922 1.010 0.944 0.908 0.979 0.927 0.954 1.001 0.991 0.940 0.919 0.716 0.747 0.916 0.947 0.975 0.801 0.839 0.886 0.858 0.894 0.884 0.872 0.866 0.835 0.861 0.842 Con.c (ug/ml) µg/mg 762.308 693.846 760.000 752.308 601.538 697.692 691.538 737.692 712.308 656.154 646.923 693.077 740.000 694.615 762.308 711.538 683.846 738.462 698.462 719.231 755.385 747.692 708.462 692.308 536.154 560.000 690.000 713.846 735.385 601.538 630.769 666.923 645.385 673.077 665.385 656.154 651.538 627.692 647.692 633.077 69.301 63.077 69.091 68.392 54.685 63.427 62.867 67.063 64.755 59.650 58.811 63.007 67.273 63.147 69.301 64.685 62.168 67.133 63.497 65.385 68.671 67.972 64.406 62.937 48.741 50.909 62.727 64.895 66.853 54.685 57.343 60.629 58.671 61.189 60.490 59.650 59.231 57.063 58.881 57.552 51 Percentage of sucrose (g/100g) 13.167 11.985 13.127 12.994 10.390 12.051 11.945 12.742 12.303 11.334 11.174 11.971 12.782 11.998 13.167 12.290 11.812 12.755 12.064 12.423 13.048 12.915 12.237 11.958 9.261 9.673 11.918 12.330 12.702 10.390 10.895 11.520 11.148 11.626 11.493 11.334 11.254 10.842 11.187 10.935 Ordinal Entry 231 232 233 234 235 236 237 238 239 240 241 242 243 244 245 246 247 248 249 250 251 252 253 254 255 256 257 258 259 260 261 262 263 264 265 266 267 268 269 270 PE 593 PE 597 PE 599 PE 604 PE 605 PE 608 PE 609 PE 610 PE 612 PE 613 PE 615 PE 616 PE 617 PE 618 PE 620 PE 621 PE 622 PE 623 PE 624 PE 625 PE 626 PE 629 PE 630 PE 632 PE 634 PE 635 PE 637 PE 639 PE 640 PE 644 PE 645 PE 646 PE 648 PE 649 PE 650 PE 659 PE 663 PE 664 PE 671 PE 674 Absorbance at 510 nm 0.804 0.809 0.823 0.813 0.755 0.898 0.813 0.779 0.734 0.865 0.768 0.783 0.823 0.822 0.778 0.803 0.805 0.812 0.821 0.786 0.713 0.711 0.862 0.733 0.663 0.924 0.820 0.878 0.847 0.791 0.862 0.870 0.795 0.816 0.999 0.751 0.630 0.738 0.762 0.757 Con.c (ug/ml) µg/mg 603.846 607.692 618.462 610.769 566.154 676.154 610.769 584.615 550.000 650.769 576.154 587.692 618.462 617.692 583.846 603.077 604.615 610.000 616.923 590.000 533.846 532.308 648.462 549.231 495.385 696.154 616.154 660.769 636.923 593.846 648.462 654.615 596.923 613.077 753.846 563.077 470.000 553.077 571.538 567.692 54.895 55.245 56.224 55.524 51.469 61.469 55.524 53.147 50.000 59.161 52.378 53.427 56.224 56.154 53.077 54.825 54.965 55.455 56.084 53.636 48.531 48.392 58.951 49.930 45.035 63.287 56.014 60.070 57.902 53.986 58.951 59.510 54.266 55.734 68.531 51.189 42.727 50.280 51.958 51.608 52 Percentage of sucrose (g/100g) 10.430 10.497 10.683 10.550 9.779 11.679 10.550 10.098 9.500 11.241 9.952 10.151 10.683 10.669 10.085 10.417 10.443 10.536 10.656 10.191 9.221 9.194 11.201 9.487 8.557 12.024 10.643 11.413 11.001 10.257 11.201 11.307 10.310 10.590 13.021 9.726 8.118 9.553 9.872 9.806 Ordinal 271 272 273 274 275 276 277 278 279 280 281 282 283 284 285 286 287 288 289 290 291 292 293 294 295 296 297 298 299 300 301 302 303 304 305 306 307 308 309 310 Entry PE 676 PE 783 PE 785* PE 787 PE 788 PE 793* PE 796 PE 802 PE 803* PE 805 PE 808 PE 809 PE 810 PE 811 PE 812 PE 813* PE 814* PE 815 PE 816 PE 817 PE 818 PE 820 PE 821 PE 823 PE 824 PE 825 PE 826 PE 827 PE 828 PE 829 PE 830 PE 831* PE 832 PE 833 PE 834 PE 835 PE 836 PE 837 PE 838 PE 839* Absorbance at 510 nm 0.760 0.919 1.023 0.824 0.918 1.036 0.869 0.906 0.995 0.757 0.859 0.908 0.970 0.849 0.935 1.044 0.999 0.786 0.934 0.969 0.879 0.950 0.736 0.893 0.920 0.890 0.896 0.955 0.927 0.952 0.943 1.048 0.888 0.819 0.916 0.815 0.975 0.899 0.808 1.021 Con.c (ug/ml) 570.000 692.308 772.308 619.231 691.538 782.308 653.846 682.308 750.769 567.692 646.154 683.846 731.538 638.462 704.615 788.462 753.846 590.000 703.846 730.769 661.538 716.154 551.538 672.308 693.077 670.000 674.615 720.000 698.462 717.692 710.769 791.538 668.462 615.385 690.000 612.308 735.385 676.923 606.923 770.769 53 µg/mg 51.818 62.937 70.210 56.294 62.867 71.119 59.441 62.028 68.252 51.608 58.741 62.168 66.503 58.042 64.056 71.678 68.531 53.636 63.986 66.434 60.140 65.105 50.140 61.119 63.007 60.909 61.329 65.455 63.497 65.245 64.615 71.958 60.769 55.944 62.727 55.664 66.853 61.538 55.175 70.070 Percentage of sucrose (g/100g) 9.845 11.958 13.340 10.696 11.945 13.513 11.294 11.785 12.968 9.806 11.161 11.812 12.636 11.028 12.171 13.619 13.021 10.191 12.157 12.622 11.427 12.370 9.527 11.613 11.971 11.573 11.652 12.436 12.064 12.397 12.277 13.672 11.546 10.629 11.918 10.576 12.702 11.692 10.483 13.313 World soybean production has increased by 76% from 6.5 billion bushels in 2000 to 11.3 billion bushels in 2014 Brazil and Argentina increased soybean production by 138% and 98%, respectively, during the same time period The United States increased soybean production by 44% between 2000 and 2014 while soybean production increased by 137% in the ROW region Most of that increase took place in other South American countries (Richard and Won, 2015) The world soybean production in the year of 2016 was approximately 313, 26 million metric tons The USA is the largest soybean with producer with 106, 93 million metric tons The other major production countries such as Brazil, Argentina and Paraguay contributed about 97, 56.5 and 8 million metric tons, respectively (USDA, 2016) Soybeans increased 24.75 USD/BU or 2.18% from 1159.25 on Friday June 17 to 1134.50 in the previous trading session Soybeans gained 182.8 USD/BU or 18.71 percent during the last 12 months from 976.50 USD/BU in June of 2015 (http://www.tradingeconomics.com/commodity/soybeans, 2016) Soybean are used for food, pharmaceutical, cosmetic, bio-fuel industries It is rich in protein carbohydrates, saponins, phytic acid and oil but has lower levels of calcium It has wide range of therapeutic as well as pharmaceutical applications It has sedative, anti-spasmodic, diaphoretic, anti-pyretic properties, with hormonal balancing effects, can be used to prevent breast cancer, prostate cancer, endometrial cancer and baldness and has great benefit to the liver and circulation It acts as a component of easy release jelly and also as an agent that provides good hardness and disintegrating property In tofu, soymilk, natto, and many other soy food products, desirable sugars including glucose, fructose, and sucrose contribute to the favorable sweet taste and are ready-todigest, while raffinose and stachyose are indigestible and cause undesirable flatulence and diarrhea (Rackis, 1975) In China, Japan, and Korea, soybeans are also used in industrial products, including oils, soap, cosmetics, resins, plastics, inks, crayons, solvents, and clothing Ordinal 351 352 353 354 355 356 357 358 359 360 361 362 363 364 365 366 367 368 369 370 371 372 373 374 375 376 377 378 379 380 381 382 383 384 385 386 387 388 389 390 Entry PE 929 PE 932 PE 934 PE 935 PE 936 PE 938 PE 939 PE 940 PE 941 PE 944 PE 947 PE 951* PE 962 PE 956 PE 958 PE 959 PE 960 PE 962 PE 963 PE 964 PE 967 PE 971 PE 973 PE 974 PE 982 PE 983 PE 984 PE 985 PE 990 PE 991 PE 993 PE 1159 PE 1160 PE 1164 PE 1165 PE 1166 PE 1170 PE 1172 PE 1175 PE 1176 Absorbance Con.c (ug/ml) at 510 nm 0.838 630.000 0.870 654.615 0.947 713.846 0.914 688.462 0.848 637.692 0.866 651.538 0.861 647.692 0.952 717.692 0.919 692.308 0.854 642.308 0.925 696.923 1.006 759.231 0.915 689.231 0.686 513.077 0.918 691.538 0.952 717.692 0.733 549.231 0.484 357.692 0.808 606.923 0.588 437.692 0.664 496.154 0.698 522.308 0.731 547.692 0.841 632.308 0.907 683.077 0.784 588.462 0.751 563.077 0.785 589.231 0.885 666.154 0.717 536.923 0.886 666.923 0.921 693.846 0.915 689.231 0.865 650.769 0.874 657.692 0.892 671.538 0.823 618.462 0.893 672.308 0.827 621.538 0.891 670.769 55 µg/mg 57.273 59.510 64.895 62.587 57.972 59.231 58.881 65.245 62.937 58.392 63.357 69.021 62.657 46.643 62.867 65.245 49.930 32.517 55.175 39.790 45.105 47.483 49.790 57.483 62.098 53.497 51.189 53.566 60.559 48.811 60.629 63.077 62.657 59.161 59.790 61.049 56.224 61.119 56.503 60.979 Percentage of sucrose (g/100g) 10.882 11.307 12.330 11.892 11.015 11.254 11.187 12.397 11.958 11.094 12.038 13.114 11.905 8.862 11.945 12.397 9.487 6.178 10.483 7.560 8.570 9.022 9.460 10.922 11.799 10.164 9.726 10.178 11.506 9.274 11.520 11.985 11.905 11.241 11.360 11.599 10.683 11.613 10.736 11.586 Ordinal Entry 391 392 393 394 395 396 397 398 399 400 401 402 403 404 405 406 407 408 409 410 411 412 413 414 415 416 417 418 419 420 421 422 423 424 425 426 427 428 429 430 PE 1179 PE 1180 PE 1183 PE 1185 PE 1187 PE 1188 PE 1192 PE 1193 PE 1194 PE 1196 PE 1197 PE 1198 PE 1199 PE 1200 PE 1202 PE 1204 PE 1205 PE 1207 PE 1208 PE 1209 PE 1210 PE 1212 PE 1213 PE 1214 PE 1215 PE 1216 PE 1217 PE 1218 PE 1219 PE 1220 PE 1221 PE 1221 PE 1225 PE 1227 PE 1228 PE 1229 PE 1235 PE 1237 PE 1241 PE 1242 Absorbance at 510 nm 0.887 0.812 0.863 0.784 0.836 0.834 0.819 0.817 0.870 0.829 0.774 0.803 0.899 0.597 0.828 0.614 0.775 0.861 0.770 0.748 0.790 0.854 0.823 0.848 0.767 0.846 0.788 0.891 0.803 0.755 0.698 0.772 0.776 0.837 0.908 0.754 0.789 0.836 0.746 0.881 Con.c (ug/ml) 667.692 610.000 649.231 588.462 628.462 626.923 615.385 613.846 654.615 623.077 580.769 603.077 676.923 444.615 622.308 457.692 581.538 647.692 577.692 560.769 593.077 642.308 618.462 637.692 575.385 636.154 591.538 670.769 603.077 566.154 522.308 579.231 582.308 629.231 683.846 565.385 592.308 628.462 559.231 663.077 56 µg/mg 60.699 55.455 59.021 53.497 57.133 56.993 55.944 55.804 59.510 56.643 52.797 54.825 61.538 40.420 56.573 41.608 52.867 58.881 52.517 50.979 53.916 58.392 56.224 57.972 52.308 57.832 53.776 60.979 54.825 51.469 47.483 52.657 52.937 57.203 62.168 51.399 53.846 57.133 50.839 60.280 Percentage of sucrose (g/100g) 11.533 10.536 11.214 10.164 10.855 10.829 10.629 10.603 11.307 10.762 10.031 10.417 11.692 7.680 10.749 7.906 10.045 11.187 9.978 9.686 10.244 11.094 10.683 11.015 9.938 10.988 10.217 11.586 10.417 9.779 9.022 10.005 10.058 10.869 11.812 9.766 10.231 10.855 9.659 11.453 Ordinal Entry 430 431 432 433 434 435 436 437 438 439 440 441 442 443 444 445 446 447 448 449 450 451 452 453 454 455 456 457 458 459 460 461 462 463 464 465 466 467 468 469 470 PE 1242 PE 1244* PE 1245 PE 1246 PE 1248* PE 1252 PE 1254 PE 1256* PE 1259 PE 1260 PE 1263 PE 1265 PE 1294 PE 1295 PE 1299 PE 1300* PE 1301* PE 1304* PE 1306 PE 1307 PE 1308 PE 1309 PE 1310 PE 1314 PE 1315 PE 1317 PE 1318 PE 1321 PE 1322 PE 1323 PE 1326 PE 1327 PE 1329 PE 1331 PE 1332 PE 1333 PE 1334 PE 1335 PE 1337 PE 1338 PE 1340 Absorbance Con.c (ug/ml) at 510 nm 0.881 663.077 1.007 760.000 0.815 612.308 0.926 697.692 1.012 763.846 0.868 653.077 0.879 661.538 0.962 725.385 0.875 658.462 0.935 704.615 0.938 706.923 1.014 765.385 0.970 731.538 0.972 733.077 0.975 735.385 1.019 769.231 0.991 747.692 1.011 763.077 0.849 638.462 0.874 657.692 0.840 631.538 0.912 686.923 0.887 667.692 0.900 677.692 0.820 616.154 0.765 573.846 0.922 694.615 0.796 597.692 0.833 626.154 0.802 602.308 0.863 649.231 0.810 608.462 0.889 669.231 0.867 652.308 0.874 657.692 0.830 623.846 0.935 704.615 0.936 705.385 0.946 713.077 0.952 717.692 0.851 640.000 57 µg/mg 60.280 69.091 55.664 63.427 69.441 59.371 60.140 65.944 59.860 64.056 64.266 69.580 66.503 66.643 66.853 69.930 67.972 69.371 58.042 59.790 57.413 62.448 60.699 61.608 56.014 52.168 63.147 54.336 56.923 54.755 59.021 55.315 60.839 59.301 59.790 56.713 64.056 64.126 64.825 65.245 58.182 Percentage of sucrose (g/100g) 11.453 13.127 10.576 12.051 13.194 11.280 11.427 12.529 11.373 12.171 12.210 13.220 12.636 12.662 12.702 13.287 12.915 13.180 11.028 11.360 10.908 11.865 11.533 11.706 10.643 9.912 11.998 10.324 10.815 10.403 11.214 10.510 11.559 11.267 11.360 10.776 12.171 12.184 12.317 12.397 11.055 Ordinal Entry 471 472 473 474 475 476 477 478 479 480 481 482 483 484 485 486 487 488 489 490 491 492 493 494 495 496 497 498 499 500 501 502 503 504 505 506 507 508 509 510 PE 1341 PE 1342 PE 1344 PE 1348 PE 1349 PE 1350 PE 1351 PE 1352 PE 1352 PE 1354 PE 1355 PE 1356* PE 1358 PE 1360 PE 1364 PE 1366 PE 1367 PE 1370 PE 1371 PE 1372* PE 1373 PE 1375 PE 1376 PE 1377 PE 1378 PE 1379 PE 1380 PE 1384 PE 1386* PE 1388 PE 1390 PE 1392 PE 1394 PE 1395 PE 1399 PE 1441 PE 1445 PE 1446 PE 1448 PE 1449 Absorbance Con.c (ug/ml) at 510 nm 0.955 720.000 0.868 653.077 0.874 657.692 0.954 719.231 0.862 648.462 0.874 657.692 0.971 732.308 0.939 707.692 0.900 677.692 0.949 715.385 0.717 536.923 1.031 778.462 0.917 690.769 0.924 696.154 0.916 690.000 0.861 647.692 0.889 669.231 0.910 685.385 0.827 621.538 1.003 756.923 0.883 664.615 0.933 703.077 0.801 601.538 0.981 740.000 0.963 726.154 0.835 627.692 0.917 690.769 0.906 682.308 1.010 762.308 0.985 743.077 0.958 722.308 0.921 693.846 0.874 657.692 0.882 663.846 0.894 673.077 0.912 686.923 0.943 710.769 0.907 683.077 0.875 658.462 0.859 646.154 58 µg/mg 65.455 59.371 59.790 65.385 58.951 59.790 66.573 64.336 61.608 65.035 48.811 70.769 62.797 63.287 62.727 58.881 60.839 62.308 56.503 68.811 60.420 63.916 54.685 67.273 66.014 57.063 62.797 62.028 69.301 67.552 65.664 63.077 59.790 60.350 61.189 62.448 64.615 62.098 59.860 58.741 Percentage of sucrose (g/100g) 12.436 11.280 11.360 12.423 11.201 11.360 12.649 12.224 11.706 12.357 9.274 13.446 11.931 12.024 11.918 11.187 11.559 11.838 10.736 13.074 11.480 12.144 10.390 12.782 12.543 10.842 11.931 11.785 13.167 12.835 12.476 11.985 11.360 11.466 11.626 11.865 12.277 11.799 11.373 11.161 Soybean oil is the primary source of biodiesel in the United States, accounting for 80% of domestic biodiesel production Soybeans have also been used since 2001 as fermenting stock in the manufacture of a brand of vodka Soybean and soybean products are a common part of the diet (https://en.wikipedia.org/wiki/Soybean, 2016) The FDA granted the following health claim for soy: "25 grams of soy protein a day, as part of a diet low in saturated fat and cholesterol, may reduce the risk of heart disease” (Henkel and John 2000) Soybean helped to reduce cholesterols and prevent and manage certain types of cancer, kidney disease, osteoporosis, diabetes, and obesity (Illinois Center for Soy Foods, 2007) In addition, one of most important agronomic characteristic for soybean and other legume species was their ability to took nitrogen from the air and convert it to metabolizable ammonium N, a process known as nitrogen fixation It also helps to keep the production costs for soybean relatively lower compared to other cops The ability of soybean plant to fix N2 has played a very significant role in maintaining the nitrogen balance in biosphere and environmental cleanness Saving billion of dollar for provided fertilizer nitrogen (Liu, 1997) 1.2 The sucrose Soybean seed are normally valued not only for their high oil contents and protein but also contain significant amount of carbohydrate, also known as saccharides with the general chemical formula Cn(H2O)m and their derivatives It including simple sugar (mono and disaccharide), oligosaccharide, and polysaccharide Recent studies have shown that some beneficial effects of dietary oligosaccharides dietary for humans’ health and there are including: Increasing population of indigenous bifid bacteria in the colon which, by their antagonistic effect, suppress the activity of putrefactive bacteria Reducing toxic metabolites and detrimental enzyme Preventing pathogenic and autogenous diarrhea by the same mechanisms as described in the reduction of detrimental bacteria Ordinal Entry 551 552 553 554 555 556 557 558 559 560 561 562 563 564 565 566 567 568 569 570 571 572 573 574 575 576 577 578 579 580 581 582 583 584 585 586 587 588 589 590 PE 1652 PE 1653 PE 1655 PE 1657 PE 1669 PE 1676 PE 1690 PE 1696 PE 1697 PE 1699 PE 1701 PE 1702 PE 1712 PE 1722 PE 1731 PE 1751 PE 1754 PE 1756 PE 1757 PE 1760 PE 1765 PE 1768 PE 1769 PE 1774 PE 1783 PE 1786 PE 1789 PE 1790 PE 1792 PE 1793 PE 1795 PE 1798 PE 1799 PE 1800 PE 1804 PE 1806 PE 1807 PE 1808 PE 1809 PE 1810 Absorbance at 510 nm 0.737 0.837 0.863 0.787 0.760 0.870 0.866 0.873 0.978 0.946 1.013 0.676 0.885 0.975 0.994 0.889 0.919 0.892 0.911 0.899 0.859 0.868 0.891 0.872 0.811 0.876 0.844 0.950 0.895 0.853 0.910 0.856 0.889 0.939 0.960 0.910 0.889 0.896 0.807 0.805 Con.c (ug/ml) 552.308 629.231 649.231 590.769 570.000 654.615 651.538 656.923 737.692 713.077 764.615 505.385 666.154 735.385 750.000 669.231 692.308 671.154 686.154 676.538 645.769 653.077 670.385 655.769 609.231 658.846 634.231 716.154 673.846 641.538 685.000 643.462 668.846 707.308 723.846 685.385 669.231 674.231 605.769 604.231 60 µg/mg 50.210 57.203 59.021 53.706 51.818 59.510 59.231 59.720 67.063 64.825 69.510 45.944 60.559 66.853 68.182 60.839 62.937 61.014 62.378 61.503 58.706 59.371 60.944 59.615 55.385 59.895 57.657 65.105 61.259 58.322 62.273 58.497 60.804 64.301 65.804 62.308 60.839 61.294 55.070 54.930 Percentage of sucrose (g/100g) 9.540 10.869 11.214 10.204 9.845 11.307 11.254 11.347 12.742 12.317 13.207 8.729 11.506 12.702 12.955 11.559 11.958 11.593 11.852 11.686 11.154 11.280 11.579 11.327 10.523 11.380 10.955 12.370 11.639 11.081 11.832 11.114 11.553 12.217 12.503 11.838 11.559 11.646 10.463 10.437 Ordinal Entry 591 592 593 594 595 596 597 598 599 600 601 602 603 604 605 606 607 608 609 610 611 612 613 614 615 616 617 618 619 620 621 622 623 624 625 626 627 628 629 630 PE 1812 PE 1817 PE 1824 PE 1825 PE 1826 PE 1827 PE 1828 PE 1832 PE 1838 PE 1846 PE 1847 PE 1852 PE 1853 PE 1855 PE 1856 PE 1869 PE 1875 PE 1878 PE 1888 PE 1892 PE 1897 PE 1898 PE 1906 PE 1908 PE 1911 PE 1912 PE 1918 PE 1922 PE 1929 PE 1931 PE 1935 PE 1936 PE 1937 PE 1938 PE 1939 PE 1941 PE 1943 PE 1950 PE 1951 PE 1954 Absorbance Con.c (ug/ml) at 510 nm 0.814 611.154 0.923 695.385 0.965 727.308 0.778 583.462 0.876 658.846 0.824 619.231 0.813 610.385 0.760 569.615 0.909 684.231 0.887 667.308 0.831 624.231 0.808 606.923 0.878 660.385 0.902 679.231 0.803 603.077 0.984 742.308 0.891 670.769 0.872 656.154 0.894 673.077 0.881 662.692 0.837 628.846 0.877 659.615 0.935 704.615 0.906 681.923 0.862 648.462 0.813 610.769 0.857 644.615 0.815 612.308 0.866 651.538 0.894 673.077 0.802 602.308 0.827 621.538 0.846 636.154 0.809 607.692 0.908 683.846 0.905 681.538 0.806 605.385 0.940 708.462 0.808 606.923 0.941 709.231 61 µg/mg 55.559 63.217 66.119 53.042 59.895 56.294 55.490 51.783 62.203 60.664 56.748 55.175 60.035 61.748 54.825 67.483 60.979 59.650 61.189 60.245 57.168 59.965 64.056 61.993 58.951 55.524 58.601 55.664 59.231 61.189 54.755 56.503 57.832 55.245 62.168 61.958 55.035 64.406 55.175 64.476 Percentage of sucrose (g/100g) 10.556 12.011 12.563 10.078 11.380 10.696 10.543 9.839 11.819 11.526 10.782 10.483 11.407 11.732 10.417 12.822 11.586 11.334 11.626 11.447 10.862 11.393 12.171 11.779 11.201 10.550 11.134 10.576 11.254 11.626 10.403 10.736 10.988 10.497 11.812 11.772 10.457 12.237 10.483 12.250 Ordinal Entry 631 632 633 634 635 636 637 638 639 640 641 642 643 644 645 646 647 648 649 650 651 652 653 654 655 656 657 658 659 660 661 662 663 664 665 666 667 668 669 670 PE 1958 PE 1962 PE 1963 PE 1964 PE 1966 PE 1969 PE 1971 PE 1973 PE 1974 PE 1975 PE 1975 PE 1977 PE 1979 PE 1983 PE 1985 PE 1987 PE 1989 PE 1990 PE 1991 PE 1993 PE 1996 PE 1998 PE 1999 PE 2001 PE 2002 PE 2005 PE 2007 PE 2008 PE 2011 PE 2012 PE 2013 PE 2194 PE 2195 PE 2205 PE 2209 PE 2217 PE 2219 PE 2228 PE 2231 PE 2232 Absorbance at 510 nm 0.958 0.855 0.971 0.951 0.933 0.952 0.941 0.929 0.857 0.872 0.888 0.853 0.852 0.858 1.015 0.908 0.912 0.871 0.813 0.879 0.962 0.963 0.926 0.950 0.982 0.888 0.828 0.865 0.942 0.869 0.937 0.927 0.866 0.892 0.855 0.872 0.921 0.806 0.859 0.829 Con.c (ug/ml) µg/mg 722.308 643.077 732.308 716.923 703.077 717.692 709.231 700.000 644.615 656.154 668.462 641.538 640.769 645.385 766.154 683.846 686.923 655.385 610.769 661.538 725.385 726.154 697.692 716.154 740.769 668.462 622.308 650.769 710.000 653.846 706.154 698.462 651.538 671.538 643.077 655.769 693.462 605.385 646.154 623.077 65.664 58.462 66.573 65.175 63.916 65.245 64.476 63.636 58.601 59.650 60.769 58.322 58.252 58.671 69.650 62.168 62.448 59.580 55.524 60.140 65.944 66.014 63.427 65.105 67.343 60.769 56.573 59.161 64.545 59.441 64.196 63.497 59.231 61.049 58.462 59.615 63.042 55.035 58.741 56.643 62 Percentage of sucrose (g/100g) 12.476 11.108 12.649 12.383 12.144 12.397 12.250 12.091 11.134 11.334 11.546 11.081 11.068 11.148 13.234 11.812 11.865 11.320 10.550 11.427 12.529 12.543 12.051 12.370 12.795 11.546 10.749 11.241 12.264 11.294 12.197 12.064 11.254 11.599 11.108 11.327 11.978 10.457 11.161 10.762 Ordinal 671 672 673 674 675 676 677 678 679 680 681 682 683 684 685 686 687 688 689 690 691 692 693 694 695 696 697 698 699 700 701 702 703 704 705 706 707 708 709 710 Entry PE 2233 PE 2240 PE 2246 PE 2258 PE 2259 PE 2263 PE 2274 PE 2296 PE 2300 PE 2307 PE 2314 PE 2315 PE 2327 PE 2329 PE 2332 PE 2334 PE 2342 PE 2343 PE 2352 PE 2354 PE 2360 PE 2361 PE 2362 PE 2363 PE 2365 PE 2369 PE 2377 PE 2378 PE 2382 PE 2233 PE 2240 PE 2246 PE 2258 PE 2259 PE 2263 PE 2274 PE 2296 PE 2300 PE 2307 PE 2314 Absorbance Con.c (ug/ml) at 510 nm 0.824 618.846 0.868 652.692 0.871 655.000 0.869 653.846 0.832 625.000 0.904 680.769 0.882 663.846 0.930 700.769 0.887 667.308 0.826 620.385 0.832 625.000 0.737 551.923 0.834 626.923 0.852 640.385 0.858 645.000 0.859 646.154 0.903 679.615 0.803 603.077 0.816 613.077 0.799 600.000 0.805 604.615 0.848 637.692 0.871 655.385 0.851 640.000 0.852 640.769 0.843 633.846 0.851 639.615 0.838 630.000 0.784 588.462 0.825 620.311 0.824 619.274 0.823 618.237 0.821 617.200 0.820 616.164 0.819 615.127 0.817 614.090 0.816 613.053 0.815 612.017 0.813 610.980 0.812 609.943 63 µg/mg 56.259 59.336 59.545 59.441 56.818 61.888 60.350 63.706 60.664 56.399 56.818 50.175 56.993 58.217 58.636 58.741 61.783 54.825 55.734 54.545 54.965 57.972 59.580 58.182 58.252 57.622 58.147 57.273 53.497 56.392 56.298 56.203 56.109 56.015 55.921 55.826 55.732 55.638 55.544 55.449 Percentage of sucrose (g/100g) 10.689 11.274 11.314 11.294 10.795 11.759 11.466 12.104 11.526 10.716 10.795 9.533 10.829 11.061 11.141 11.161 11.739 10.417 10.590 10.364 10.443 11.015 11.320 11.055 11.068 10.948 11.048 10.882 10.164 10.714 10.697 10.679 10.661 10.643 10.625 10.607 10.589 10.571 10.553 10.535 ... study, a total of 710 mutant soybean lines from a mutation population derived from EMS treatment were analyzed by the GOPOD/invertase method We obtained 390 soybean lines with greater sugar content... 1974) Also, the application of rapid analytical techniques such as near-infrared reflectance spectroscopy (NIRS) has many advantages compared to standard and traditional techniques such as chromatographic... preliminary analysis, as a results we obtained 390 lines with sucrose content of larger than that of Pungsanamul The remaining 320 lines were lower Selected 31 soybean lines have larger than that sucrose