Growth-related Molecular Markers Screening Of Large Yellow Croaker Based On The Dart Technology

Large yellow croaker (Larimichthys crocea) is the major marine economic fish of China, mainly distributingin coast of Zhejiang, Fujian, Guangdong Province. In the past two decades, due to overfishing, wild large yellowcroaker had already become extinct in china. To protect fish resource and meet the demand of market, largeyellow croaker farming was gradually developed. But because of years of farming activities without breeding,farmed large yellow croaker appeared slow growth, precocious puberty, low immunity and other phenomena,therefore the urgent need for genetic improvement of cultured large yellow croaker. However, large yellowcroaker’s economic traits, such as growth, sex determination, disease resistance, stress resistance in most casesshowed quantitative genetic. The Traditional breeding method is based on phenotypic selection, cycle is long, lowefficiency, difficult to accurately obtain the needed economic traits, so it is necessary to using molecular markertechnology from gene level, using economic traits related molecular marker assisted selection an breeding, toimprove breeding efficiency. Growth as one of the most important economic traits, can directly reduce the activityof breeding resources investment, enhance the economic benefits. This study built a large yellow croaker genomiclibrary, and using bulked segregant analysis and DArT technology to screen molecular markers related to growthtraits in large yellow croaker. The main results are as follows:Using the Large yellow croaker which named "Donghai No.1" as the experimental material, the length of199individuals measured. The maximum is16.30cm. The minimum is11.50cm, the mean is13.45cm, thestandard deviation is1.3367, kurtosis is0.192, partial degrees is0.210, Shapiro-Willie normal distribution test p=0.257, accord with normal distribution (p>0.05). Established a map of normal distribution base on body length,body length of individuals wihich is in10%high value recorded as "extremely big group", the10%low valuerecorded as "extreme" small group ", for preliminary screening DArT marker related to growth traits.Using7kinds of genomic DNA complexity reduction method (PstI/AluI, PstI/BanII, PstI/Bsp1286I,PstI/BstNI, PstI/HaeIII, PstI/RsaI, PstI/TaqI) to construct7genomic libraries of large yellow croaker. The librarystorage capacity were8×105,1×106,4×106,1.2×106,4×106,4×106,1.2×106, the average insert size is886bp,691bp,820bp,607bp,623bp,765bp,721bp. Therefore, the construction of the libraries meet the requirements offurther experiments.820clones from each genomic libraries were used as templates to make probe for high-density chip. Andpolymorphism rate of7kinds of large yellow croaker genomic DNA complexity reduction method is validatedThe results from high to low is PstI/RsaI17.66%, PstI/TaqI16.83%, PstI/BstNI14.14%, PstI/BanII13.96%,PstI/HaeIII13.02%, PstI/Bsp1286I8.10%, PstI/AluI6.92%, validation data showed that PstI/RsaI reduced thecomplexity of the genome of large yellow croaker had the best effect, PstI/TaqI followed.3360clones, randomly selecting from PstI/RsaI genomic libraries, were used as templates to make probe forscreening chip."Extreme big group" and "extreme small group "was hybridized, The fluorescence value analysedby using fuzzy C-means clustering analysis method (fuzzy degree is1.5) and ANOVA to get0or1markers, andconducte a two element matrix.The results show that clustering relationship is consistent with the grouping.18candidate polymorphism fragments from the matrix were screened, as candidate DArT markers related to growth traits in large yellow croaker. These results lay the foundation for further verification of DArT marker relatedgrowth traits.This study provided theoretical support for the experimental biology genetic breeding of large yellow croaker,had important theoretical and practical significance to the birth of large yellow croaker groups with good growthtraits.