The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Human subjects: Ethical approvalThis study was performed in five different avenues: Clinical Embryology Laboratory at IVIRMAValencia (Spain), University of Cambridge (United Kingdom), National Research Center forAssisted Reproductive Technology and Reproductive Genetics (China), California Institute ofTechnology (United States), and Foundation for Embryonic Competence New Jersey (UnitedStates). All the work complies with The International Society for Stem Cell Research (ISSCR)guidelines.Experiments performed in Spain: The work in the Clinical Embryology Laboratory at IVIRMAValencia was approved by the National Commission of Human Reproduction (CNRHA), theGeneral direction of research, innovation, technology and quality and the ethics committee ofClinical Research IVI Valencia, which complies with Spanish law on assisted reproductivetechnologies (14/2006). A total of 260 excess and cryopreserved donated day 3 humanembryos from 95 IVF patients were used. Embryos were not created for research. Couplesdonating their embryos were informed of the objectives of the project, the nature of theexperiments, and the conditions of the research. No financial inducements were offered fordonation. The average age of women was 28.12 {plus minus} 4. The fixed embryos were analysed at theUniversity of Cambridge. This work was evaluated by the Human Biology Research EthicsCommittee of the University of Cambridge (reference HBREC.2017.27).Experiments performed in the United Kingdom: The work performed at the University ofCambridge was in accordance with the Human Fertility and Embryology Authority (HFEA)regulations (license reference R0193). Ethical approval was obtained from the "HumanBiology Research Ethics Committee" of the University of Cambridge (referenceHBREC.2017.21). Informed consent was obtained from all patients from CARE Fertility Groupand Herts & Essex fertility clinics, who donated excess and cryopreserved embryos used inthis study after completing their IVF treatment. Embryos were not created for research. Priorto giving consent, patients were informed about the specific objectives of the project, and theconditions that apply within the license, offered counselling and did not receive any financialinducements. In this study, we used 15 donated embryos at the 2 pronuclei stage (day 1 d.p.f.),which were warmed and cultured in the Cambridge Laboratory according to the aboveregulations.Experiments performed in China: The work performed in the National Research Center forAssisted Reproductive Technology and Reproductive Genetics was conducted under theregulations of the Human Biomedical Research Ethics Guidelines (regulated by NationalHealth Commission of the People's Republic of China on 1 December 2016) and the HumanEmbryonic Stem Cell Research Ethics Guidelines (regulated by China National Center forBiotechnology Development on 24 December 2003). These regulations and guidelines allowexcess 3PN human gametes, and/or human embryos created or genetically manipulated invitro and those cultured for no more than 14 days, to be used for scientific researches. Theaim and protocols involved in this study were reviewed and approved by the InstitutionalReview Board (IRB) of Reproductive Medicine, Shandong University. The protocols includethe use of 3PN embryos and the siRNA injections. All embryos were donated after informedconsent was obtained. The embryos were excess and fresh 3PN zygotes generated from invitro fertilisation, with the donor females age ranging from 22 to 40 years old. These embryoswere used for microinjection experiments. Embryos were not created for research. Patientswere informed about the objectives, experimental approach and potential outcomes of theresearch prior to the donation. They were offered counselling to aid them in their decision.Experiments performed in California (United States): The work at California Institute ofTechnology was approved by the California Institute of Technology Committee for theProtection of Human Subjects (IRB number 19-0948). Human embryos at the zygote stagewere obtained from the University of Southern California (USC) through the pre-existing USCIRB-approved Biospecimen Repository for Reproductive Research (HS-15-00859) afterappropriate approval was obtained unanimously from the Biorepository Ethics Committee. AtUSC Fertility, excess cryopreserved embryos were donated after completion of IVF; embryoswere not created for research purposes. Patients were informed of the general conditions ofthe donation, objectives, and methodology of human embryo research. They were offeredcounselling and alternatives, including discarding embryos and continued cryopreservation.Patients were informed that they would not benefit directly from donation of embryos toresearch. A total of 41 donated human embryos at the zygote 2 pronuclei stage (day 1 postfertilisation.) from 4 IVF patients were used. The average age of women was 44. The embryoswere warmed at USC Fertility per usual IVF procedure and then transferred to the CaliforniaInstitute of Technology for the remainder of the protocol.Experiments performed in New Jersey (United States): the research was approved by theWestern IRB (Clinical IRB 20031397). Embryos used in this study were excess fresh embryosto the IVF treatment of IVI-RMA patients. Informed consent was obtained from all the couplesthat donated their embryos. They were informed of the general objective, methodology andpotential outcomes of the research. Patients did not receive any financial compensation forthe donation. Embryos were not created for research.

© 2021, Zhu et al.

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