Method Of Cell Culture From Testes And Cryopreservation For Spermatophore Of Litopenaeus Vannamei

Litopenaeus vannamei farming is an important part of fishery economy in our country. Studying on excellent varieties breeding and gender controlling to reduce disease, increase production and reduce the production cost, is an important development direction of the c farming.Spermatogonial stem cells (SSCs) are one kind of multifunctional diploid cells in the seminiferous epithelium of the testes tissue on shrimp. This experiment that culturing the testis tissue cells of Litopenaeus vannamei, could be a fundamental study of spermatogenesis mechanism model, gene controlling, sperm screening, gender controlling, etc.. In this experiment, a vannamei shrimp was dissected in an aseptic condition. The cells was obtained from the testis tissue after being moved out, cut into minced and percolated by the sieve, then incubated at28℃degree and90%humidity in medium199supplemented with lmol/L HEPES buffer solution,20%fetal bovine serum and100IU/ml double-antibiotic consisted of penicillin and streptomycin. Cells could be observed by inverted microscope and recorded by photo shotting; Cells could be identified by hematoxylin-eosin staining and recorded by photo shotting. In this experiment, the photo of cells cultured from testes of Litopenaeus vannamei and stained cells was shotted, and this experiment was completed basicly.Spermatophore is one unique reproductive structure of decapod animal including Litopenaeus vannamei for protecting and transmitting the sperm. This study on cryopreservation for spermatophore of Litopenaeus vannamei is a fundamental study of variety breeding for vannamei including sperm screening, artificial insemination etc., and benefit to germplasm preservation, genetic diversity protection, genetic breeding and production of Litopenaeus vannamei. In this experiment, the spermatophore of Litopenaeus vannamei was collected into an EP tube with a mixture of cultivate layer consisted of liquid-state rat tail tendon type. I,2×medium1640, fetal bovine serum and matrigel, then EP tube was sealed and stored at4℃. Survival rate of sperm from the spermatophore could be detected by eosin-nigrosine staining. In this experiment, the survival rate of sperm from the spermatophore of Litopenaeus vannamei after cryopreservation was53.15±12.35%.