黄萎病菌毒素联合法鉴定棉花对黄萎病的抗性

【目的】黄萎病菌是危害棉花最严重的病菌，又属于被检疫的病菌，田间接菌鉴定棉花抗病性受到检疫的限制，通过比较多种鉴定棉花抗黄萎病的方法，提出一种使用黄萎病菌毒素进行联合鉴定的方法，以替代传统病圃鉴定法。【方法】将待测棉花品系种植于人工气候室内，以常规管理模式培养棉苗，在21 d后将所种植棉苗的根部放入黄萎病菌毒素中浸泡，浸泡72 h时统计病情指数；在待测棉花品系的盛花期取棉株倒数第三片叶片，用打孔器在叶片上取10-20个直径约1.5 cm的叶圆盘，将所取叶圆盘浸泡于黄萎病菌的毒素当中，经过24 h后观察所浸泡的叶圆盘黄化程度，并进行统计分析；综合上述两种方法的鉴定结果，对待测棉花品系的黄萎病抗性作出评价。【结果】通过对多种鉴定方法的比较，毒素蘸根法、无底塑钵菌液浇根法和菌液蘸根法所需的鉴定时间较长，而毒素浸根法和毒素浸泡叶圆盘法所需鉴定时间较短；在应用毒素浸根法进行鉴定时，最适毒素浓度为15 μg·mL^-1，在此浓度下，毒素浸根法的鉴定结果与田间病圃法鉴定结果的相关系数为0.94（P＜0.01）；在使用毒素浸泡叶圆盘法进行鉴定时，浸泡叶圆盘的最适毒素浓度为18 μg·mL^-1；在应用毒素浸泡叶圆盘法鉴定时，应于盛花期选取待测棉花的倒三叶的叶圆盘进行鉴定，其鉴定结果与常规病圃法鉴定结果的相关系数为0.92（P＜0.01）；2013年和2014年两年的试验结果证明联用毒素浸根法和毒素浸泡叶圆盘法的鉴定结果与常规病圃法的鉴定结果相关系数较高。2013年其与常规病圃法的鉴定结果相关系数为0.94（P＜0.01），2014年为0.95（P＜0.01）。【结论】通过联用毒素浸根法和毒素浸泡叶圆盘法可以准确、环保、便捷地鉴定棉花抗黄萎病性，可以在一定程度上代替传统的病圃鉴定法，其结果比用单一鉴定方法更为准确可靠。

A Combined Identification Method for the Verticillium Wilt Resistance in Cotton by Using Pathogen Toxin

【Objective】 The Verticillium dahliae is the most harmful fungus for cotton plants and belongs to quarantine fungus. It is restricted to use Verticillium fungus to identify the Verticillium wilt resistance of cotton in the field. Some identification methods were compared in this research. In order to replace this traditional identification method, a new combined method by using Verticillium toxin instead of using its fungus was found in this paper. And this research provided an accurate, environment-friendly, convenient method to breed the cotton varieties with resistance to Verticillium wilt. 【Method】Method Ⅰ: The cotton was sowed in the greenhouse with conventional management and their roots were soaked in the toxin of Verticillium after 21 days. Then the disease indexes were counted after 72 h. Method Ⅱ: The antepenultimate leaves were collected at full-bloom stage of cotton, and they were cut to about 10-20 small leaf disc (the diameter is 1.5 cm) by hole puncher. Whereafter it was put into the Verticillium toxin, and counted the disease indexes and yellowing degree after 24 h. The final results were combined with two methods to evaluate the Verticillium resistance of candidate cotton lines. 【Result】This paper suggested that it would spend longer time in identifying the Verticillium wilt resistance by using dipping the root with toxin, sprinkling the spore suspension on root in non-bottom plastic film bottle and dipping the root with spore suspension. However, it would spend shorter time in identifying the Verticillium wilt resistance by using soaking the leaf disc with toxin and soaking the root with toxin. The best concentration of toxin in this research was 15 μg·mL^-1, when soaking the root with toxin was used to identify the Verticillium wilt resistance. Compared with the conventional method, the correlation coefficient was 0.94 (P＜0.01) in that concentration. When soaking the leaf disc with toxin was used to identify the Verticillium wilt resistance, the best concentration of toxin in this research was 18 μg·mL^-1. The antepenultimate leaves should be collected at full-bloom stage of cotton and the correlation coefficient was 0.92 (P＜0.01) compared with the conventional method. The combined method was a quick, accurate and environment-friendly method to identify the Verticillium wilt resistance of cotton. Compared with the conventional method, the correlation coefficient of combined approach was higher through the experimental results in 2013 and 2014. In 2013 and 2014, the correlation coefficient was 0.94 (P＜0.01) and 0.95 (P＜0.01), respectively.【Conclusion】 Using the combined method which includes soaking the leaf disc with toxin and soaking the root with toxin can identify the Verticillium wilt resistance accurately, environment-friendly and conveniently. It can replace the conventional method by using this combined method. The combined identification method was more reliable than the single identification method in identification.