Embryo Transfer Technology Research Articles

Production, cryopreservation, and transfer of Murrah embryos through Multiple Ovulation and Embryo Transfer (MOET) technology in Sri Lanka

Production, cryopreservation, and transfer of Murrah embryos through Multiple Ovulation and Embryo Transfer (MOET) technology in Sri Lanka

Development of a biotechnology process for the production of a novel hyperglycosylated long-acting recombinant bovine follicle-stimulating hormone.

Follicle-stimulating hormone (FSH) is an important protein used for bovine ovarian hyperstimulation in multiple ovulation and embryo transfer technology (MOET). Several attempts to produce bovine FSH (bFSH) in recombinant systems have been reported, nonetheless, up to date, the most commonly used products are partially purified preparations derived from porcine or ovine (pFSH or oFSH) pituitaries. Here we describe the development of a biotechnology process to produce a novel, hyperglycosylated, long-acting recombinant bFSH (LA-rbFSH) by fusing copies of a highly O-glycosylated peptide. LA-rbFSH and a nonmodified version (rbFSH) were produced in suspension CHO cell cultures and purified by IMAC with high purity levels (>99%). LA-rbFSH presented a higher glycosylation degree and sialic acid content than rbFSH. It also demonstrated a notable improvement in pharmacokinetic properties after administration to rats, including a higher concentration in plasma and a significant (seven-fold) reduction in apparent clearance (CLapp). In addition, the in vivo specific bioactivity of LA-rbFSH in rats was 2.4-fold higher compared to rbFSH. These results postulate this new molecule as an attractive substitute for commercially available porcine pituitary-derived products.

Bovine superovulation and embryo transfer – how to make babies!

Bovine embryo transfer technology is the pursuit of managing genetics for a more preferred calving outcome. This technology has allowed for great genetic advancements by increasing selec­tion pressure or decreasing generation interval. Embryos can be derived by 2 methods. In-vivo embryo collection allows the donor dam to grow and develop the embryo. In-vitro produced embryos require fertilization and development of the embryo in a lab setting. This is a discussion of the concepts of embryo production and transfer. These methods have not changed drastically over the years; however, their application continues to evolve. Embryo transfer provides the opportunity to make more of the desired bovine babies.

Effects of donor age and reproductive history on developmental potential of ovum pickup oocytes in Japanese Black cattle (Wagyu)

The objectives of this study were to analyze the (1) effects of donor age and multiparity on development of in vitro fertilization (IVF) embryos after ovum pickup (OPU), (2) effects of repeated and consecutive OPU-IVF procedures on embryo development, and (3) embryo production from OPU-IVF in donors with differing embryo yields after multiple ovulation and embryo transfer technology (MOET) in Japanese Black cattle (Wagyu). Donors were pre-treated with low-dosage follicle-stimulating hormone (FSH; 200 IU total), and oocytes were collected via OPU and fertilized by IVF to generate blastocysts. The number of oocytes collected per OPU session per donor was lower in heifers (2–4 years old, 5.3 oocytes) than in primiparous and pluriparous cows (2–10 years old, 13.6–19.1 oocytes; P < 0.05). Rates of blastocyst development for oocytes from heifers (33.1%) were lower than for those from cows (2–10 years old, 44.1–54.3%; P < 0.05), and average blastocyst yield/OPU/animal was lower in heifers (3.7) than in 5–6 years old cows (10.1; P < 0.05). Donors undergoing five consecutive OPU-IVF sessions after low-dosage FSH showed similar oocyte retrieval (12.2–15.1 oocytes per OPU/animal), blastocyst development rates (35.6–45.0%), and embryo yield/OPU/animal (4.8–5.8; P > 0.05) across sessions. Additionally, embryo yield from OPU-IVF was significantly improved in animals with previous low embryo yield from MOET (5.9 vs. 2.6, respectively, P < 0.05). These results indicate that Wagyu cows with previous births can be more productive as OPU-IVF donors than heifers, and oocytes from donors undergoing to five consecutive OPU-IVF cycles are competent for embryo development without loss of embryo yield/OPU/animal. Moreover, OPU-IVF can be used for embryo production and breeding from all elite Japanese Black cattle, regardless of previous low embryo yield in routine MOET.

The Poor Performance of Neu5Gc for Endometrial Receptivity in Embryo Transfer

Neu5Gc is a common sialic acid in mammals, but humans cannot synthesize it independently. When Neu5Gc in the diet enters the body, the body produces antibodies against Neu5Gc, which in turn promotes the immune response and chronic inflammation and may be closely related to embryo transfer rejection. Therefore, Neu5Gc has gained widespread interest from gynecologists, nutritionists, and immunologists. The same problem occurs during embryo transfer, where the embryo carries paternal gene products and is a hapten relative to the maternal endometrium. Successful implantation requires a receptive endometrial environment. Moreover, with the widespread application of in vitro fertilization and embryo transfer technology, nonhuman amino acids may be introduced into the embryo culture medium, mainly the genetic engineering-derived essential serum albumin in embryo culture medium, which may introduce the nonhuman sialic acid Neu5Gc and cause embryo transfer rejection.

Evolution over the last 40 years of the assisted reproductive technologies in cattle - the Brazilian perspective for embryo transfer and resynchronization programs (part II).

The productivity of the beef and dairy industries depends directly on the reproductive efficiency and genetic gain of the herd, which are directly associated with the appropriate use of Assisted Reproductive Technologies (ARTs). The objective of this review is to show from a Brazilian perspective the evolution over the last 40 years of ARTs related to ovulation resynchronization programs and embryo transfer in cattle. Despite significant improvements and high fertility obtained in timed artificial insemination (TAI) protocols (Sales et al., 2024 - Part I), the improvement of the use of in vitro-produced embryos, development of resynchronization programs, and the advance in Doppler ultrasonography (Doppler-US) for reproductive assessments of bovine females were the ARTs that presented the greatest relevance on reproductive effectiveness in cattle. In the last seven years, the embryo transfer (ET) technology using in vitro-produced (IVP) embryos took over the conventional ET of in vivo produced embryos after donor's superovulation. Also, procedures and pregnancy rates after ET of IVP embryos were improved in dairy and beef operations. The Doppler-US allows the identification of non-pregnant females at an early stage based on the evaluation of blood perfusion of the corpus luteum. Recent studies in beef and dairy cows indicate satisfactory accuracy when Doppler-US is used at 20-22 days after TAI. Consequently, super-early resynchronization programs have been developed and are being implemented in commercial programs, thereby facilitating earlier conception through the use of semen from superior bulls, providing genetic and economic improvements in herds. Likewise, the assessment of luteal function by Doppler-US allows the selection of embryo recipients with greater receptivity, and consequently may increase the effectiveness of timed ET programs.

Current state of horse embryo transfer technology in Russia and factors influencing its effectiveness

The article presents data on activity in the field of embryo transfer in the Russian horse breeding system and analyzes of the factors influencing the success of the use of this technology. The results of Russian reproductive specialists work over the past 15 years have been collected (144 procedures of embryo recovery in 52 mares). The influence of age and degree of synchronization of ovulations between donors and recipients on the level of embryo recovery and survival rates was analyzed. It has been established that the highest embryo recovery rate is observed in mares of 2- 10 years old (yo) - 84.6%. In mares of 11-16 yo, this figure decreased to 64.3% (p˂ 0.05), and for 17-24 yo mares - to 42.8% (p˂ 0.001). The survival rate of embryos in the uterus of recipient mares of 3-14 yo remains at 70- 71%, in mares of 15-20 yo it sharply decreased to 42.8%. It has been shown that embryo transfer to a recipient whose ovulation occurred 1 day earlier or within a period of up to 5 days after the donor's ovulation provides a survival rate of 79% until the 35th day of pregnancy. When the recipient ovulation occurs 2-4 days earlier than the donor ovulation, the result of embryo transfer was dramatically reduced (7 ,7±7,4%, p˂0.001).

14 Production and transfer of bovine embryos through the hub-and-spoke model: an innovative model to promote embryo transfer technology

14 Production and transfer of bovine embryos through the hub-and-spoke model: an innovative model to promote embryo transfer technology

Effect of different manual puncture methods on donkey embryo before vitrification

The application of embryo recovery and transfer technology in the donkey industry is far lower than that of horses and cattle. Sometimes the recovered embryos could not be transferred in time, which required embryo cryopreservation. The embryo cryopreservation technology is more conducive to the preservation and transportation of recovered embryos with excellent genetic traits. However, this technique for donkey embryos is not efficient and needs further optimization. The objective of this study was to evaluate the effect of different manual puncture methods on the viability and pregnancy rates of vitrified-thawed donkey embryos. A total of 117 donkey embryos were recovered on day 7–8 post-ovulation and were divided into four groups by random assortment. There were 28 embryos without puncture or cryopreservation (Control). 58 embryos were manually punctured using a 29G needle (VG, n = 29) or microneedle with a sharp tip of <10 μm (VM, n = 29), then vitrified in 15% EG + 15% DMSO + 0.5 M sucrose. Another 31 embryos were punctured with a microneedle and vitrified with 10% EG + 10% DMSO +0.5 M sucrose +2 mol/L proline (VMP). Both fresh embryos and vitrified-thawed embryos were incubated for 3 h (38.5 °C, 5% CO2 in air) before embryo transfer. The results showed that the embryo recovery rates on day 7.5 and 8 were higher than day 7 (P < 0.05). After incubation, dead cells were assessed and the percentages of dead cells in VM and VMP were lower than that in VG (P < 0.05), although both were higher than those in Control (P < 0.05). The pregnancy rates after 23 days post transfer were assessed and the results showed that the pregnancy rate in VG (8.0%) was lower than that in Control (41.7%), VM (24.0%) and VMP (29.6%) (P < 0.05). No pregnancies resulted from the 10 embryos with diameters ≤650 μm in VG, which lower than either VM (33.3%) or VMP (38.9%) (P < 0.05). While, there was no difference in pregnancy rates among all vitrification groups when embryos were >650 μm in diameter (P > 0.05). In conclusion, the embryo recovery rate on day 7 after ovulation was relatively low, and it was more appropriate to extend it to day 8. Microneedle puncture could reduce embryo damage and achieve a higher pregnancy rate compared with 29G needles. Proline has the potential to improve donkey embryo cryopreservation.

CRISPR/Cas mediated disruption of BMPR-1B gene and introduction of FecB mutation into the Caprine embryos using Easi-CRISPR strategy

Bone Morphogenetic Proteins play a significant role in ovarian physiology and contribute to the reproductive fitness of mammals. The BMPR-1B/FecB mutation, a loss of function mutation increases litter size by 1–2 with each number of mutated alleles in sheep. Considering demand-supply gap of the meat industry, and low replacement rate of indigenous caprine species, the conservative BMPR-1B locus can be explored, and FecB mutated goats can be produced. The experiment one produced CRISPR/Cas mediated KO transferable caprine embryos, and experiment two generated caprine embryos with desired FecB mutation using Easi-CRISPR strategy. In the KO experiment, Cas9 and BMPR-1B guide RNA (100:100ng/ul) were electroporated into single stage caprine zygotes at 750V, 10 ms and 1pulse using Neon transfection system. In the second experiment, phosphorothioate (PS) modified single-stranded oligodeoxynucleotide (ssODN) was used as an HDR template along with CRISPR components (100:100ng/ul, ssODN 100ng/ul). The precise time and method of electroporation, RNP format of CRISPR components and PS modified asymmetric ssODN were the factors that affected the production of mosaicism free BMPR-1B edited caprine embryos. The editing efficiency of KO and KI experiments was 68.52 and 63.16% respectively, and successful production of goats with higher mean ovulation rate can be realized with addition of embryo transfer technology to these experiments.

Biotechnologies for accelerated reproduction of Kalmyk cattle

In Russia, the number of beef cattle until recently was concentrated mainly in Kalmykia and adjacent regions. However, the lack of purposeful breeding has led to the fact that the purebred livestock of Kalmyk meat breed has significantly decreased and lost its unique qualities. Introduction of embryo transfer biotechnology can solve the problem of restoring breed qualities under the condition of accelerated reproduction of genotyped livestock. The aim of the research was to study Kalmyk breed response to polyovulation considering further increase in transferrable embryo productivity - the main indicator specific to each breed. As the formation of the breed took place in pasture conditions with a pronounced seasonal sexual cycle, there were doubts about the possibility of using embryo transfer technology in any season. The experiments were carried out on Kalmyk cows at Kalmytskoye farm in 2010 and at the Regional Research and Production Center for Reproduction of Farm Animals, Kalmyk State University, in 2022. The results of the study showed that it was better to synchronize sexual hunting using prostaglandins after 3-4 months after calving (May-October). According to the results of immunogenetic testing, 10 breeding cows and 2 breeding bulls were selected as donors. In September-October, in order to test the embryo transfer method in nomadic animal husbandry, the first work was carried out to stimulate poliovulation and obtain embryos from 3 donor cows. A total of 23 embryos and eggs were obtained, 10 of which were suitable for transplantation. Transferrable embryos were frozen on a ZEM-4 software freezer after equilibration in 1.5 M ethylene glycol cryoprotector solution.

IETS management of the challenges associated with embryo pathogen interaction

One of the very first challenges the International Embryo Transfer Society (IETS) addressed was concern about disease transmission via the transfer of in vivo–derived (IVD) bovine embryos. IETS commissioned its Import/Export Committee, later named Health and Safety Advisory Committee (HASAC), to resolve this matter, with the assistance of the Data Retrieval Committee following its formation in 1991. Since its first meeting in 1984, considerable achievements have been made, including meeting the numerous challenges created by the many innovations in this industry. Based on research studies and their designs, the IETS HASAC developed a system for categorising pathogens and diseases potentially susceptible to interaction with IVD embryos. This has been instrumental in defining safe operating protocols and ultimately leading to the development of the now universally accepted techniques for certification of embryo health. The close cooperation of IETS/HASAC with the World Organization of Animal Health (WOAH, formerly OIE) has facilitated the establishment of guidelines for regulators worldwide, thus ensuring the safety of international trade with embryos, while avoiding unjustified regulatory measures. In addition, IETS/HASAC produced and published the IETS Manual: A Procedural Guide and General Information for the Use of Embryo Transfer Technology Emphasising Sanitary Procedures for the embryo transfer industry (1st edition, 1987; 5th edition, 2023). This manual and its updates were designed to provide the industry world-wide with a source of information on safe and sanitary handling procedures for embryos, to describe the procedures necessary to ensure that the transfer of embryos does not result in transmission of pathogenic agents or disease, and to ensure consistent and accurate identification of embryos. The result of these 40 years of IETS/HASAC involvement is that embryo transfer technology is recognised as having a comparative advantage in international movement of germplasm.

Historical Background and Significance of Embryo Transfer Technology in Cattle with its Relevant Applications

Historical Background and Significance of Embryo Transfer Technology in Cattle with its Relevant Applications

TINGKAT KEBERHASILAN TRANSFER EMBRIO (TE) BERDASARKAN KEGIATAN SELEKSI RESIPIEN DAN PELAKSANAAN TE DARI TAHUN 2018 SAMPAI DENGAN 2021 DI KABUPATEN ROKAN HULU

Implementation of the application of embryo transfer technology (ET), requires a program that can meet the wishes of the livestock community in terms of improving the genetic quality of liverstock quickly. The program is related to the proposal of beef cattle by field officers, selection of recipient cattele by the embryo transfer team which includes: exterior assessment, such as the Body Condition Score (BSC), interior assessment such as rectal palpation of the internal genitals (cervix, uterus an ovaries). The next activity is the implementation of embryo transfer such as how handle recipient cattel; wheather the recipient has been in Artificial Insemination (AI) (twinning program) or single ET, epidural anastesi, thowing frozen embryos in water at 38℃, insertion of embryo straw into the ET gun/plastic ET, rectal palpation, cervical examination, insertion of the ET gun, embryo deposition to the uterine horn, recording the results of ET, examination of pregnancy results of ET at the age of 2 months, monitoring of recipients who are already pregnant until elivery, reporting the results of single ET briths and ET twinning(AI/ET).&#x0D; This technical report is the result of the embryo transfer (ET) program activities that took place from 2012,2018,2019,2020,and 2021, which in this technical report discusses the success rate of embryo transfer (ET) based on recipient selection and impelemtation activities embryo transfer (ET).

COMPARISON OF SEMEN QULAITY FOR THREE LINES OF HOLSTEIN BULLS: 1. SOME IMMEDIATE AND MICROSCOPIC CHARACTERISTICS

This study was conducted to compare the semen immediate and microscopic characteristics of the three lines of Holstein bulls. Twenty-one Holstein bulls were divided into three groups belonging to the three lines of Holstein bulls; born in Iraq (Australian origin; n = 8), resulting from embryo transfer technology (New Zealand origin; n = 7), as well as the first generation (F1) resulted from two parents obtained from embryo transfer technology in Iraq (New Zealand origin; n = 6). The ET and L groups showed greater (P≤0.01) ejaculate volume, live sperms percentage, and plasma membrane integrity percentage compared to the F1 group of bulls. Moreover, ET bulls exhibited higher (P≤0.01) sperm concentration than L and F1 bulls. The ET and F1 bulls were superior (P≤0.01) to L bulls in the percentages of sperm’s cell individual motility and normal sperms. The percentage of DNA damage was significantly (P≤0.05) decreased in the F1 group compared to the L group but did not differ from those of the ET group. In conclusion, the ET and F1 Holstein bulls were superior to L bulls in most immediate and microscopic semen characteristics and their adaptation to the Iraqi environment.

Evaluation of the In Vitro Embryo Production of Farm Animals Under the Circumstances of Embryo Transfer Technology

“Embryo transfer” technology is one of the deterministic techniques related to the production of in vitro or in vivo produced embryos. This technique controls to a large extent the integrity of the developmental component of early embryos and their ability to develop later until birth. Pregnancy rates resulting from the transferred embryos were widely dispersed and associated with many variables and factors. The developmental stage of early embryos had a significant impact on pregnancy rates, as most studies agreed to transfer embryos in the blastocyst and morula stages due to the high rates of pregnancy (40%-100%) compared to the blastomeres. Embryo cryopreservation technology (rapid and slow) competed to a large extent with fresh embryos in pregnancy rates, even surpassing the latter in some cases, as the percentage approached 100%. The resulting pregnancy rates varied greatly (up to 95%) according to the method of transferring and hormonal induction. The results were mainly based on the size and type of the animal on the one hand and on the nature of the biological activity and the specific function of the hormones (progesterone, prostaglandin F2α, gonadotropin - releasing hormone and follicle - stimulating hormone, etc.) involved in regulating estrus in donors and recipients on the other hand. This review concluded that the embryo transfer technology has given many scenarios that cannot lead to an inevitable result in judging the efficacy of the technology of in vitro embryo production, but the results are encouraging and require further efforts.

Improving genetic quality of local livestock for achieving food security

Indonesia has local livestock breeds, both ruminant and non-ruminant. The existence and preservation of those animals is very important. The government, the concerned parties and farmers of community member are required to be responsible for maintaining the productivity of livestock. The quality genetic could be improved by applying breeding and selection. There are three efforts can be used to improve the livestock population and productivity, those are artificial insemination, embryo transfer, and separation technology of spermatozoa X and Y. The objective to be achieved is to help meet the need of food, especially animal protein, of the domestic community, in addition to the welfare of the community.

Multiple ovulation and embryo transfer in sheep: Effects of embryo developmental stage and quality on viability in vivo under farm conditions.

Multiple ovulation and embryo transfer (MOET) technologies are integral to genetic improvement programs in the sheep industries. Despite the protocols being well established, previous findings regarding the effects of embryo properties on MOET success remain contradictory. The objective of this study was to determine the effects of embryo developmental stage and quality on embryo viability following transfer to recipient ewes. Data including details of 377 embryos collected from 45 Merino donor ewes were obtained from MOET trials conducted on three separate farms on day 6 after laparoscopic artificial insemination (AI). A total of 270 embryos were classified as being of transferrable grade (grade 1: n = 233; grade 2: n = 37). One or two transferrable grade embryos were transferred to each of 256 synchronised recipient ewes and pregnancy diagnosis was performed on day 36 after embryo transfer. Embryos at the hatched blastocyst stage tended to have greater viability in vivo compared to embryos at the late morula stage (59.0 ± 10.6% vs. 36.2 ± 9.7%; P = 0.083). The viability of grade 1 embryos was greater than that of grade 2 embryos (53.6 ± 7.8% vs. 35.9 ± 10.2%; P < 0.05). The results suggest that the success of the MOET trials was influenced by the transfer of embryos at the late morula stage, almost half of which were classified as grade 2 embryos. These findings highlight the importance of following strict embryo quality grading criteria to inform the most economical management of recipient ewes and maximize pregnancy outcomes.

The Criminal Law Regulation of Surrogacy Accessory Crime

Abstract With the continuous improvement of artificial reproduction technology, in vitro fertilization (IVF) or embryo transfer technology has developed rapidly. At the same time, the proportion of crimes related to surrogacy is also increasing. Surrogacy has different problems in judicial practice, laws and regulations, and theory. Specifically, it can be subdivided into: due to the lack of surrogacy regulation in current laws and regulations, there are related problems centered on surrogacy behavior. On the one hand, it often encounters bottlenecks due to the lack of legal support in judicial practice; On the other hand, at the theoretical level, because surrogacy involves the exclusive rights of life, such as physical rights and reproductive rights, this practical issue is controversial and has mixed opinions, which leads to the fault phenomenon in the application of this issue at the judicial level. Based on this situation, this paper starts from the practical level, enumerates the typical cases involving criminal acts in the process of surrogacy, and regulates them with the existing criminal law norms. Secondly, refine the characteristics and elements of surrogacy behavior in the above-mentioned practical level, think about this phenomenon, and redefine the concept of surrogacy based on the current social environment. Finally, on the basis of the above concept of surrogacy, return to the level of criminal law, take the dichotomy of surrogacy regulation as the guidance, and take the surrogacy institution as the main body of crime. It jointly explores the way to solve the black industrial chain of surrogacy.

Production of Agu piglets after transfer of embryos produced in vitro.

The present study was conducted to examine the feasibility of in vitro embryo production and transfer technologies for producing piglets of Agu, an Okinawan indigenous pig breed. After collection of oocytes from surgically dissected ovaries, they were subjected to in vitro maturation. After in vitro maturation/fertilization, a total of 616 putative embryos were transferred into four commercial Western pig recipients, one of which became pregnant and farrowed a total of eight Agu piglets. These results demonstrate that in vitro embryo production using ovaries from Agu females is useful for breeding management and conservation of indigenous breeds.