Methods for the production and regeneration of Lactobacillus casei protoplasts are described. Protoplasts of L. casei strains were obtained by treatment with mutanolysin or with mutanolysin and lysozyme together in a protoplast formation buffer containing 0.02 M HEPES (N-2-hydroxyethylpiperazine-N′-2-ethanesulfonic acid) (pH 7.0), 1 mM MgCl2, 0.5% gelatin, and 0.3 M raffinose. Cells were regenerated on a complex medium supplemented with bovine serum albumin, MgCl2, CaCl2, gelatin, and raffinose. Lengthy digestion with lytic enzymes inhibited the capacity of protoplasts to regenerate. The optimum conditions of protoplast formation varied from strain to strain. Using predetermined optimal conditions it was possible to prepare protoplasts of several L. casei strains and regenerate them with 10 to 40% efficiency. The methods were applicable to other species of lactobacilli as well.
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机译：描述了生产和再生干酪乳杆菌原生质体的方法。干酪乳杆菌菌株的原生质体是通过在含有0.02 M HEPES（N-2-羟乙基哌嗪-N'-2-乙磺酸）（pH 7.0），1 mM MgCl2的原生质体形成缓冲液中用变溶菌素或变溶菌酶和溶菌酶一起处理而获得的，0.5％的明胶和0.3 M的棉子糖。在补充了牛血清白蛋白，MgCl2，CaCl2，明胶和棉子糖的复杂培养基上再生细胞。用裂解酶长时间消化会抑制原生质体再生的能力。原生质体形成的最佳条件因菌株而异。使用预定的最佳条件，可以制备几种干酪乳杆菌菌株的原生质体，并以10％至40％的效率对其进行再生。该方法也适用于其他种类的乳杆菌。
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