Establishment and application of GISH reaction system in triploid loach hybrid
ZHOU He， ZHUANG Zi-xin， ZHA Huang-yuan， LIANG Yu-ting， XU Qi-zheng，ZHANG Rui， LIN Zhong-qiao， LI Ya-juan
Key Laboratory of Marine Bio-resources Sustainable Utilization in Liaoning Province's University，Key Laboratory of Mariculture＆ Stock Enhancement in North China's Sea， Ministry of Agriculture， Dalian Ocean University， Dalian 116023， China
Abstract: In this study， reaction system of genomic in situ hybridization(GISH)as a powerful means to identify hybrids was optimized under several experimental parameters and was used to evaluate hybridization between Paramisgurnus dabryanus(2n＝48)and natural tetraploid loach Misgurnus anguilicaudatus(4n＝100)to study mechanism of natural tetraploid loach and to realize industrial production of tetraploid loach.The results showed that the probe DNA had 500 bp to 1000 bp in length at 105 ℃ for 2 min；the blocking DNA had 100 bp to 500 bp in length at 110℃for 10 min in the GISH reaction system of triploid hybrid loach.The screening of probe/blocking DNA ratio showed better hybridization signal at both 1∶25 and 1∶50.GISH analysis of reciprocal cross offspring by whole genome DNA of loach as probe DNA and by the whole genome DNA of natural tetraploid loach as blocking DNA revealed that twenty-four chromosomes from parental loach showed hybridization signal at the centromeres.No hybridization signal points was observed in fifty chromosomes from parental tetraploid loach.The findings offered most intuitive evidence for identification of triploid hybrid loach offsprings and provided molecular genetics basis for further provement of the tetraploid loach in China as the heritable tetraploid with four genomes，which could produce gamete of 2n，guiding breeding of triploid loach.
2018, Vol. 33 
