Chlorella sorokinus starch-deficient mutant strain and its application
A technology for synthesizing starch and chlorella, which can be used in mutant preparation, single-cell algae, biofuel, etc., can solve the problems that are difficult to be widely used, and achieve the effects of reducing the risk of disease, shortening the time required for induction and cultivation, and requiring no equipment
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Embodiment 1
[0056] Embodiment 1: Mutagenesis screening of mutant strains
[0057] 1. Determine the germination rate and semi-lethal rate of Chlorella under the solid plate of heterotrophic medium
[0058] Determination of germination rate: smear 100 μL of algae liquid on each Endo solid medium, the number of cells per 100 μL is 1000, 500, and 200, and each cell concentration gradient is made in three parallels, and then cultured in a 37°C incubator for 5 days , to determine the cell germination rate according to the number of single colonies grown from each plate.
[0059] The formula of Endo heterotrophic medium is as follows:
[0060] Glucose 20g / L; KNO 3 2g / L; KH 2 PO 4 1.2g / L; MgSO 4 ·7H 2 O 1.2g / L; Trisodium Citrate 0.2g / L; CaCl 2 2H 2 O mother liquor (1000×) 1ml; FeSO 4 ·7H 2 O and EDTA·2Na mother solution (1000×) 1mL; trace element mother solution (1000×) 1mL.
[0061] in:
[0062] CaCl 2 Preparation of mother liquor: 105g CaCl 2 2H 2 O dissolved in 1000ml H 2 O; ...
Embodiment 2
[0078] Example 2, under the induction of high light and low nitrogen, the dry weight curve, starch and oil content of mutant strains were measured, and the best mutant strains were selected
[0079] (1) The medium formula of low-nitrogen (1 / 16N) autotrophic medium BG11 is as follows:
[0080]
[0081]
[0082] The ingredients of Trace metal mix A5 are:
[0083] Element Mother liquor concentration (g / L) h 3 BO 3
2.86 MnCl 2 .4H 2 o
1.86 ZnSO 4 .7H 2 o
0.22 Na 2 MoO 4 .2H 2 o
0.39 CuSO 4 .5H 2 o
0.08 Co(NO3) 2 .6H 2 o
0.05
[0084] K 2 HPO 4 It is sterilized separately from Ferric ammonium citrate mother solution, and added after other solutions are sterilized.
[0085] The configuration method of A5 (you need to add each ingredient to the water in turn, stir until dissolved and then add another ingredient).
[0086] Finally, adjust the pH of the prepared BG11 medium to 7.1 with 1...
Embodiment 3
[0139] Example 3: Determination of the dry weight curve, cell growth curve, oil content and TAG content of the SLM2 mutant strain with the highest oil production under heterotrophic fermentation culture
[0140] The mutant strain SLM2 with low starch content and high oil content was selected and cultured at high density in a fermenter to study its dry weight, cell growth and oil content under full medium.
[0141] Proceed as follows:
[0142] Select monoclonal algae strain SLM2 and wild type to activate in 20ml Endo liquid medium for 2 days (primary culture); when entering the logarithmic phase of growth, adjust monoclonal algae strain SLM2 and wild type to the same cell concentration and transfer to In a 250ml Erlenmeyer flask, the final volume of the algae liquid is 100ml. The temperature is 37°C, 150rpm (secondary culture); on the third day, transfer 40ml of algae liquid to a 1000ml Erlenmeyer flask, add fresh Endo medium to 400ml (tertiary culture) and cultivate for 3.5 d...
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