Acipenser dabryanus spermatogonium culture solution and application thereof
A technology of spermatogonia and Dabry's sturgeon, which is applied in the biological field and can solve the problems of the undiscovered Dabry's spermatogonia separation and culture method, etc.
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Embodiment 1
[0049] A Dabry's sturgeon spermatogonia culture fluid, comprising:
[0050]
[0051] the rest is SFM, the pH of the culture solution is 8.0.
[0052] The preparation method of described dabry's sturgeon spermatogonia culture fluid comprises the following steps:
[0053] 1. Configuration of the mother liquor of each component:
[0054] (1) Configuration of 10ml500g / LBSA mother solution (100X):
[0055] Use a balance to accurately weigh the required 0.05g of BSA with an accuracy of 0.0001g:
[0056] According to the formula: measuring weight = formula concentration × total volume of BSA mother liquor
[0057] Put the weighed BSA into the beaker, add 6ml SFM basal culture medium, fully dissolve BSA, continue to add For SFM basal culture medium, dilute the dissolved BSA solution to 10ml.
[0058] (2) Configuration of 10ml 200mmol / LL-Glutamine mother solution (100X):
[0059] Accurately weigh 0.3654gL-Glutamine with a balance, with an accuracy of 0.0001g:
[0060] Acco...
Embodiment 2-6
[0081] A Dabry's sturgeon spermatogonia culture fluid, comprising:
[0082]
[0083] The Dabry's sturgeon serum is the serum of 2-year-old male Dabry's sturgeon, and the rest are SFM medium.
Embodiment 8
[0085] A kind of Dabry's sturgeon spermatogonia culture medium is used in cultivating spermatogonia, comprising
[0086] (1) Preparation of Acipenser dabryi testis cell suspension:
[0087] The following operations were all carried out in a sterile biological safety cabinet.
[0088] 1) Move the cleaned testis tissue of Acipenser dabfieldii into a petri dish, cut the testis tissue into pieces, add 0.25% trypsin digestion solution at the ratio of digestion solution:tissue block=4ml:1g (v / w), and culture at 20°C Digest in the box and pipette once every 30min.
[0089] 2) After digesting the testis tissue in step 1) for 3 hours, add L-15 medium + 20% fetal bovine serum to terminate the digestion, and filter through a 40 μm filter. Centrifuge at 200g for 5min at 4°C and resuspend in L-15 medium + 20% fetal bovine serum.
[0090] 3) Cell counting, and staining with trypan blue: cell suspension=1:1 (v / v), after calculating the viable cell rate under a microscope, use L-15+20% fet...
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