| Soybean(Glycine max(Linn.)Merr.)is not only one of the most important oil crops in the world,but also the most important plant protein resourcein food and animal feed.,The consumption demand of soybean of our country increases continuously in recent years.With the loosingn of related policies on transgenic technology,improving the level of soybean transgenic technology and cultivating excellent soybean varieties have great great commercial values.However,there are many problems in the genetic transformation of soybean,such as strict requirements for manual operation to cut the stem cells of cotyledon,and higher ratio chimeras of the transformed plants,resulting in lower positive rate of plants and higher false positive rate,and difficulty in offspring screening.As a commonly used screening marker for genetic transformation plants,glufosinateammonium concentration plays a crucial role in shoot emergenceselection of transgenic plants.In this paper,we mainly studied the effect of glufosinate-ammonium concentration on the positive rate of soybean transformed plants to optimize the concentration of glufosinate-ammonium of trnsgeneic system.In this study,STTM-172 vector was selected and Williams 82 soybean was used as acceptor material.(1)Eight combinations of glufosinate-ammonium concentrations(10 μM and 30 μM)were set for soybean explants at the early stage of bud induction,the late stage of bud induction,and the bud elongation stage,respectively.(Group 1:10 μM、10 μM、10 μM;Group 2:10 μM、10 μM、30 μM;Group 3:30 μM、10 μM、10 μM;Group 4:30 μM、30 μM、30 μM;Group 5:30 μM、30 μM、10 μM;Group 6:10 μM、30 μM、30 μM;Group 7:10 μM、30 μM、10 μM;Group 8:30 μM、10 μM、10 μM)(2)The germination rate and seedling shoot emergence rate of soybean explants under each combinations were calculated.The lowest value was 82.62±0.57%;At the late stage of bud induction,the highest germination rate of soybean explants was65.29±2.52%,and the lowest germination rate was 50.33±0.12%.During shoot elongation,the highest and lowest elongation rate of soybean explants were 50.07±2.43%and 37.82±2.75%,respectively.(3)Under the eight glufosinate-ammonium concentration combinations,the sixth group(10 μM,30 μM,30 μM)showed the highest seedling shoot emergence rate(32.48±2.98%).The fourth group(30 μM,30 μM,30 μM)had the lowest shoot emergence rate(18.57±0.71%).The shoot emergence rate of the sixth group was significantly higher than that of the other groups.There was no significant difference in seedling shoot emergence rate between group 1 and 2,but it was higher than that of group5 and 7.There was no significant difference in seedling shoot emergence rate between group 5 and 7,which was lower only than group 1 and 2.There was no significant difference in seedling shoot emergence rate between groups 3 and 8,which was the lowest.Therefore,the highest shoot emergence rate was obtained with the glufosinate-ammonium concentration combination of group 6.(4)The transgenic plants were further identified by applying glufosinate-ammonium with1 μL/m L Glufosinate-ammonium spraying on leaves)and further molecular identification of exogenous fragment insertions.The sixth group also gave the highest positive rate was,which was 66.51±2.80%.the lowest positive rate(41.11±8.39%)was the fourth group.Then the positive rate of transformed plants in each group was compared by ANOVA,and the positive rate of transformed plants in the first and fourth groups was significantly lower than that in the other groups.There was no significant difference in the positive rate of transformed plants in the second,third,fifth,sixth,seventh and eighth groups.(5)Based on the analysis of shoot emergence rate and positive rate of transformed plants in each group,the highest shoot emergence rate and positive rate of transformed plants could be obtained under the treatment of the sixth group of glufosinate-ammonium concentration combination,so this combination of glufosinate-ammonium concentration was selected for transformation experiments with other vectors,and the average positive rate of transformed plants was 63.14%. |
PDF Full Text Request