FERTILIZATION IN-VITRO OF MOUSE OVA FROM INBRED AND OUTBRED STRAINS - COMPLETE PREIMPLANTATION EMBRYO DEVELOPMENT IN GLUCOSE-SUPPLEMENTED KSOM
FERTILIZATION IN-VITRO OF MOUSE OVA FROM INBRED AND OUTBRED STRAINS - COMPLETE PREIMPLANTATION EMBRYO DEVELOPMENT IN GLUCOSE-SUPPLEMENTED KSOM
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DOI：
10.1095/biolreprod53.2.431
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发表时间：
1995-08-01
影响因子：
3.6
通讯作者：
BIGGERS, JD
中科院分区：
文献类型：
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作者：
SUMMERS, MC;BHATNAGAR, PR;BIGGERS, JD
最近描述了一种源自连续单纯形优化方法 (SOM) 的新培养基，它克服了 CF1 培养小鼠品系体外胚胎中超过两个细胞的发育障碍。与之前的报道相反，葡萄糖对 SOM 中的胚胎发育至囊胚阶段没有显着的抑制作用。 SOM 的修饰（指定为 KSOM）增加了 Na+ (95 mM) 和 K+ (2.5 mM) 的浓度（已在其他地方描述过），也支持超出两细胞块的生长。 KSOM 产生更高的压实率、更大的囊胚产量以及滋养层细胞的细胞分裂率。我们通过改变 KSOM 中的葡萄糖浓度（0.2 mM 或 5.56 mM）重新检查了葡萄糖的影响，并确定了这些培养基支持 CF1 雌性 x B6D2F1 雄性受精卵在囊胚阶段植入前发育的能力。葡萄糖对囊胚期的发育没有明显的抑制作用，囊胚的产率通常为85%-90%。本研究中对 KSOM 进行了改良，命名为改良 KSOM，其中增加了葡萄糖浓度 (5.56 mM)，并补充了 4 mg ml(-1) BSA，现已证明可支持 CF1 卵子与杂种 B6D2F1/CrIBR 精子的体外高受精率，并随后使受精卵从两细胞阶段发育为高产囊胚。因此，改良的 KSOM 单一培养基支持 CF1 卵子的体外受精和完整的植入前胚胎发育，而无需更换培养基来克服双细胞块。 CF1卵子的囊胚总产率为80%-85%。原核、双细胞和早期囊胚阶段的胚胎移植实验获得了体外受精 CF1 卵母细胞的高着床率和活胎率。通过获得两种 F1 杂交品系的高体外受精率和囊胚发育率，证实了改良 KSOM 在小鼠卵子体外受精中的通用性：B6D2F1/J（C57BL/6J 雌性 x DBA/2J 雄性）和B6CBAF1/J（C57BL/6J 女 x CBA/J 男）。
A new medium derived from the use of sequential simplex optimization methods (SOM) that overcomes the block to development: beyond two cells in vitro in embryos of the CF1-cultured strain of mouse has recently been described. Contrary to previous reports, glucose was shown to have no significant inhibiting effect on embryo development to the blastocyst stage in SOM. A modification of SOM, designated KSOM, with an increased concentration of Na+ (95 mM) and K+ (2.5 mM), which has been described elsewhere, also supports growth beyond the two-cell block. KSOM produces a higher rate of compaction, a larger yield of blastocysts, and an increased rate of cell division of the trophoblast cells, We have reexamined the glucose effect by varying the concentration of glucose (either 0.2 mM or 5.56 mM) in KSOM and determined the ability of these media to support preimplantation development of CF1 female x B6D2F1 male zygotes through the blastocyst stage. Glucose is shown to have no significant inhibiting effect on development to the blastocyst stage, The yield of blastocysts is typically 85%-90%. A modification of KSOM derived from this study, designated modified KSOM, with an increased concentration of glucose (5.56 mM) and supplemented with 4 mg ml(-1) BSA is now shown to support high rates of fertilization in vitro of CF1 ova with hybrid B6D2F1/CrIBR sperm and subsequent development of zygotes beyond the two-cell stage to blastocysts in high yield. A single medium, modified KSOM, thus supports both fertilization in vitro of CF1 ova and complete preimplantation embryo development without the need for a change of medium to overcome the two-cell block. The overall yield of blastocysts from CF1 ova is 80%-85%. Embryo transfer experiments at the pronuclear, two-cell, and early blastocyst stages gave high rates of implantation and live fetuses from in vitro-fertilized CF1 oocytes, The general utility of modified KSOM for the fertilization of mouse ova in vitro is confirmed by obtaining high rates of fertilization in vitro and blastocyst development of two F1 hybrid strains: B6D2F1/J(C57BL/6J female x DBA/2J male) and B6CBAF1/J(C57BL/6J female x CBA/J male).