[The detection of living triploid of Haliotis discus hannai--directly making chromosome sample of mantle and epipedium tentacle].

Ploidy detection method of living body is a very important component part in polyploid breeding. There were few reports about chromosome ploidy detection of adult abalone (Haliotis discus hannai), except preparing adult abalone chromosome by means of chopping method using gill tissues and squashing method using gonad, also with these methods the abalone should be killed for getting these tissues. In polyploid breeding living triploid abalones are needed, so ploidy detection technique of living abalone is especially important. In this paper the orthogonal experiment of three factors and three levels L9(3(4)) using mantle and epipodium tentacle as material of living tissues which were vido immersing in PHA solution for preparing chromosome and count the frequence of cell division was first reported. Three factors and three levels were raising temperature: 16 degrees C, 18 degrees C, 20 degrees C; PHA concentration: 0.5%, 1.0%, 1.5%; sampling time: 5:00-6:00, 17:00-18:00, 23:00-24:00, respectively. The tests were repeated two times. The test results showed that the optimal patterns effecting the mitotic frequency of three factors and three levels in triploid abalone detection of chromosome from mantles and epipodium of living bodies were: raising temperature 20 degrees C, PHA concentration 1.0%, and sampling time 17:00-18:00. The order of three factors was: raising temperature-->PHA concentration-->sampling time. Under optimal pattern of three factors and three levels many clear, proper in length and well-scattered metaphase chromosomes could be obtained. The advantages of preparing chromosome using mantles and epipodium tentacle were sampling convenient, no body size restriction and no influence of survival of abalone. After using PHA treated tissues, mitotic frequency could be improved and 1.00% PHA concentration was the best. The key of chromosome preparation was the cells in mitotic metaphase, so the time of sampling was important. During 17:00-18:00, sampling of abalone many mitotic cells could be obtained because abalones were vigorously active at night. Biological zero degree of abalone was 7.6 degrees C. Under 7.6 degrees C abalone could not grow. At 20 degrees C abalones move and prey the most vigorously and grow fastest. Therefore a lot of cells are in mitotic metaphase.