Three Methods for Rapid Selection of Transgenic Cotton Plants in Laboratory with Kanamycin as Indirect Marker

Three methods were developed to rapidly identify Bt-transgenic cotton plants with kanamycin-resistant gene as indirect selective marker.The first method was to culture uncoated seeds on the kanamycin containing medium(kan-medium),the difference between transgenic and non-transgenic plants was revealed after seeds germinated on the medium.According to the color of the cotyledons,kan-resistant plants were discovered at optimal concentration of 0.75 g·L-1.The second method was to paint kanamycin solution(kan-solution) on the cotyledons directly with optimal concentration at 4.0 g·L-1,by which after 4-7 days,plants with cotyledon normal green were considered as kan-resistant plants.The third one was that a pore was dug on cotyledons and dropped kan-solution in.The best concentration of the kan-solution was 2.0 g·L-1.All the plants could be rapidly screened out in 4-7 days after kanamycin treatment by the three methods.The efficacy of above methods for transgenic plant indirect identification were tested using PCR analysis with Bt-gene specific primers,giving a examination ratio of more than 90%.Moreover,all the seeds decorticated could be more easily identified with kanamycin treatments.