MOESM1 of Perturbations in imprinted methylation from assisted reproductive technologies but not advanced maternal age in mouse preimplantation embryos
Creators: name: Kindsfather, Audrey
name: Czekalski, Megan
name: Pressimone, Catherine
name: Erisman, Margaret
name: Mann, Mellissa
Publisher: name: figshare
Year: 2019
Subjects: subject: Biochemistry
subject: Cell Biology
subject: Genetics
subject: FOS: Biological sciences
subject: FOS: Biological sciences
subject: Molecular Biology
subject: Physiology
subject: Pharmacology
subject: Developmental Biology

description: Additional file 1: Figure S1. Correlation of mouse and human ages on which we based our mouse maternal age model. Figure S2. Imprinted methylation acquisition at the normally methylated maternal Snrpn ICR in individual mouse blastocysts from mothers of increasing age. Embryos from spontaneously ovulated (A) virgin (1.5 to 2 months old), (B) young maternal age (&gt;2-6 months old), (C) middle maternal age (&gt;6-10 months old), and (D) advanced maternal age (&gt;10 months old) females (n=10-22 embryos; n=3-7 females per age group). Each box encloses embryos analyzed from one female. Each block represents an individual embryo. Each line denotes an individual strand of DNA with the maternal allele [C57BL/6(CAST7)] on the left and the paternal allele (C57BL/6) on the right. Embryo designation is at the top left of each block and percent methylation is at the top right. Black circles, methylated CpGs; white circles, unmethylated CpGs; ARTs, assisted reproductive technologies. Figure S3. Imprinted methylation acquisition at the normally methylated maternal Kcnq1ot1 ICR in individual blastocysts from mothers of increasing age. Embryos from spontaneously ovulated (A) virgin (1.5 to 2 months old), (B) young maternal age (&gt;2-6 months old), (C) middle maternal age (&gt;6-10 months old), and (D) advanced maternal age (&gt;10 months old) females (n=9-16 embryos; n=3-5 females per age group). See Figure S2 for details. Figure S4. Imprinted methylation acquisition at the normally methylated paternal H19 ICR in individual blastocysts from mothers of increasing age. Embryos from spontaneously ovulated (A) virgin (1.5 to 2 months old), (B) young maternal age (&gt;2-6 months old), (C) middle maternal age (&gt;6-10 months old), and (D) advanced maternal age (&gt;10 months old) females (n=11-21 embryos; n=3-6 females per age group). See Figure S2 for details. Figure S5. Imprinted methylation maintenance unaltered by maternal weight. (A) Positive correlation between maternal age and maternal weight. Black circles indicate female mice from which both oocytes and blastocysts were collected; white circles indicate female mice from which only oocytes were collected. (B-D) No association between maternal weight and imprinted methylation at the (B) Snrpn, (C) Kcnq1ot1, or (D) H19 ICRs. Diamonds represent the mean methylation of maternal (red) or paternal (blue) alleles for individual embryos. Figure S6. Imprinted methylation maintenance at the normally methylated maternal Snrpn ICR in individual blastocysts from virgin females after ART treatment. (A) SO only; (B) EC only with 5% O2; (C) SO+EC with 5% O2; (D) SO+EC with 21% O2 (n=10-13 embryos; n=3 females per treatment group). Each box encloses embryos analyzed from one female. Each block denotes an individual embryo. Each line denotes an individual strand of DNA with the maternal allele [C57BL/6(CAST7)] on the left and the paternal allele (C57BL/6) on the right. Embryo designation is at the top left of each block and percent methylation is at the top right. Black circles indicate methylated CpGs, white circles indicate unmethylated CpGs. SO, superovulation; EC, in vitro embryo culture. Figure S7. Imprinted methylation maintenance at the normally methylated maternal Kcnq1ot1 ICR in individual blastocysts from virgin females after ART treatment. (A) SO only; (B) EC only with 5% O2; (C) SO+EC with 5% O2; (D) SO+EC with 21% O2 (n=10-17 embryos; n=3-4 females per treatment group). See Figure S6 for details. Figure S8. Imprinted methylation maintenance at the normally methylated paternal H19 ICR in individual blastocysts from virgin females after ART treatment. (A) SO only; (B) EC only with 5% O2; (C) SO+EC with 5% O2; (D) SO+EC with 21% O2 (n=9-16 embryos; n=3 females per treatment group). See Figure S6 for details. Figure S9. At least one blastocyst from every ART-treated litter lost imprinted methylation on the normally methylated (A) Snrpn, (B) Kcnq1ot1, or (C) H19 ICRs. Diamonds represent the average percent methylation of maternal (red) or paternal (blue) alleles for individual embryos. ARTs, assisted reproductive technologies; SO and S, superovulation; EC and C, embryo culture; SC, superovulation plus embryo culture. Figure S10. Imprinted methylation maintenance at the normally methylated maternal Snrpn ICR in individual blastocysts from advanced maternal age females after ART treatment. Each box encloses embryos analyzed from one female. Each block denotes an individual embryo. Each line represents an individual strand of DNA with the maternal allele [C57BL/6(CAST7)] on the left and the paternal allele (C57BL/6) on the right. Embryo designation is at the top left of each block and percent methylation is at the top right. Black circles, methylated CpGs; white circles, unmethylated CpGs; SC, superovulation plus embryo culture. Figure S11. Imprinted methylation maintenance at the normally methylated maternal Kcnq1ot1 ICR in individual blastocysts from advanced maternal age females after ART treatment. See Figure S10 for details. Figure S12. Imprinted methylation maintenance at the normally methylated paternal H19 ICR in individual blastocysts from advanced maternal age females after ART treatment. See Figure S10 for details. Table S1. Comparison of treatment groups using Fisher exact test. Table S2. Comparison of maternal age and treatment groups using Fisher exact test. Table S3. Bisulfite mutagenesis and PCR amplification.