1. HRM analysis of CYTB PCR products from SCNT embryos.
HRM analysis of CYTB PCR products from SCNT embryos. The different fluorescence curves were plotted by LightScanner HRM analysis software (Version 2.0). The dilutions of plasmids that carry PCR fragments at the ratio of donor cell:recipient oocyte of 50:50, 30:70, 10:90, 3:97, 1:99, and 0:100 are indicated by orange, blue, red, green, purple, and pink lines, respectively. SCNT embryos at the blastocyst stage are indicated in gray (n = 5/group). Each standard and sample were analyzed in triplicate. The different curves that overlap with or are below the pink line are considered to harbor only oocyte mtDNA. (A) Embryos derived from nondepleted cells. Two out of five embryos possessed donor cell mtDNA at levels of 10 and 30%, and are indicated by the black and red arrows, respectively. (B) Embryos derived from nondepleted cells in the presence of TSA. Two out of five embryos possessed donor cell mtDNA at levels of 3%, and are indicated by the blue arrow. (C) Embryos derived from depleted cells. All embryos harbored only oocyte mtDNA. (D) Embryos derived from depleted cells in the presence of TSA. All embryos harbored only oocyte mtDNA.
Kanokwan Srirattana, and Justin C. St. John G3 2017;7:
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