Recycling of a single human blastomere fixed on a microscopic slide for sexing and diagnosis of specific mutations by various types of polymerase chain reaction  Zhi-Ying He, M.S., Hung-Ching Liu, Ph.D., Carol Ann Mele, B.A., Lucinda L Veeck, Owen Davis, M.D., Zev Rosenwaks, M.D.  Fertility and Sterility  Volume 72, Issue 2, Pages 341-348 (August 1999) DOI: 10.1016/S0015-0282(99)00241-1

FIGURE 1 Sequential PCRs on microscopic slides fixed single blastomere. A plastic ring was attached to the slide to form a well, and a disaggregated blastomere was then fixed inside the well. After fixation, washing, PCR with the first set of primers was run inside the well for 10 cycles and then removed to continue the rest of 20 cycles before identification of the amplified product. The blastomere was then washed thoroughly and made ready for the second run of PCR with the second set of specific primers and thus recycled for a total of five runs with five different sets of primers. Fertil Steril ©1999 Fertility and Sterility 1999 72, 341-348DOI: (10.1016/S0015-0282(99)00241-1)

FIGURE 2 Identification of amplified products: Seven isolated blastomeres were individually fixed on microscopic slides to perform five sequential PCRs for sexing (ZFX/ZFY), mutations of cystic fibrosis transmembrane regulator proteins (W1282X and ▴F508), human sickle cell β-globin, and steroid 21 hydroxylase. Lane M: Designed as the molecular weight markers; lanes 1–7: Results of seven studied blastomeres. Fertil Steril ©1999 Fertility and Sterility 1999 72, 341-348DOI: (10.1016/S0015-0282(99)00241-1)

FIGURE 3 The DOP-PCR was coupled with duplex or multiplex PCR. The genomic DNA of the biopsied blastomere can be amplified randomly and uniformly by DOP-PCR to have multiple gene (N) detection. The number of detected genes can be further increased if DOP-PCR was coupled with duplex or multiplex PCR to 2N or MN, respectively. Therefore, theoretically, this method can be used to detect whole genomic profiles from a single blastomere. Fertil Steril ©1999 Fertility and Sterility 1999 72, 341-348DOI: (10.1016/S0015-0282(99)00241-1)