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Sucrose pretreatment for enucleation: An efficient and non‐damage method for removing the spindle of the mouse MII oocyte
Author(s) -
Wang MinKang,
Liu JiLong,
Li GuangPeng,
Lian Li,
Chen DaYuan
Publication year - 2001
Publication title -
molecular reproduction and development
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.745
H-Index - 105
eISSN - 1098-2795
pISSN - 1040-452X
DOI - 10.1002/1098-2795(20010401)58:4<432::aid-mrd11>3.0.co;2-y
Subject(s) - oocyte , biology , enucleation , polar body , metaphase , microbiology and biotechnology , andrology , anatomy , chromosome , embryo , genetics , medicine , gene
Oocytes enucleated at metaphase II stage can support reprogramming of transferred nucleus and further developing to term. However, the first polar body in mice sometimes migrates away from the original place of expulsion, so the chromosomes of the oocyte will displace from the first polar body. Thus, it is not always possible to successfully enucleate according to the position of the first polar body. Here we use sucrose treatment to visualize metaphase spindle fibers and chromosomes with standard light microscopy. In the manipulation medium containing 3% sucrose, oocytes of poor quality become shrunken, deformed or fragmented, while oocytes of good quality in the same medium would show a swelling around the metaphase chromosomes and a transparent spindle area, shaped like “∞” and “0”. So it is easy to remove the well‐distinguished spindle and chromosomes in oocytes of good quality. Re‐examined by Hoechst 33342 stain under the UV light, the enucleation rate was 100%. There was no significant difference in IVF and cleavage rates between the sucrose treatment and the control group. In conclusion, this study demonstrated that 3% sucrose pretreatment can give a method for evaluating embryo quality and more importantly, it can, under a common microscope, allow the visualization of the spindle and chromosomes in oocytes of good quality and hence efficiently improve enucleation rate without any harm. Mol. Reprod. Dev. 58:432–436, 2001. © 2001 Wiley‐Liss, Inc.