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Background: The blastocyst-stage embryo has been considered more advantageous for increasing the cumulative live birth rate (CLBR) at fresh embryo transfer (ET) compared to the cleavage-stage embryo. However, it remains uncertain whether this advantage extends to specialized subpopulations, such as women with thin endometrium (TE), who are characteristic of impaired endometrial receptivity. Thus, this study aims to evaluate the difference in the CLBR between cleavage-stage and blastocyst-stage embryos at fresh ET specifically in women with TE. Methods: A retrospective cohort comprising 1089 women from three centers, ranging from September 2017 to January 2022, was established. These women were diagnosed with TE (defined as endometrium thickness <= 8 mm) and underwent their first fresh ET. To create a comparable cohort between the cleavage and blastocyst groups while adjusting for key covariates, the propensity score matching (PSM) method was employed. The primary outcome assessed was the CLBR per woman. Both cohorts underwent Kaplan-Meier analysis, Cox proportional hazard models, cumulative incidence function (CIF) curve analysis, and Fine-Grey competing risk models to ascertain the impact of embryo stage at fresh ET on CLBR. Additionally, a sensitivity analysis was conducted within a subgroup defining thin endometrium as an endometrium thickness (EMT) < 7 mm. Results: In the matched cohort after PSM, the CLBR was comparable between groups (p=0.331). However, the cleavage-stage fresh ET was associated with an elevated risk of low birth weight (LBW) (p=0.005) and small for gestational age (SGA) (p=0.037). Kaplan-Meier analysis showed that the median number of embryo transfer cycles was 2 in the cleavage group and 3 in the blastocyst group. The CLBR for the cleavage group reached 78.1%, while the blastocyst group reached 60.0% after 5 cycles of embryo transfers (log-rank test, p=0.09). A multivariable Cox proportional hazard model indicated no significant association between the embryo stage at fresh ET and CLBR (HR=0.80, 95% CI=0.60-1.07). The CIF curve and Fine-Grey competing risk models demonstrated similar results. These analyses were repeated in the original cohort before PSM and in the subgroup with EMT < 7 mm, and the results remained robust. Conclusion: For TE women receiving fresh ET, the choice between the cleavage-stage embryo and the blastocyst-stage embryo yields comparable CLBR. However, selecting the cleavage-stage embryo is associated with increased risks of LBW and SGA births. [ABSTRACT FROM AUTHOR]

BackgroundThe blastocyst-stage embryo has been considered more advantageous for increasing the cumulative live birth rate (CLBR) at fresh embryo transfer (ET) compared to the cleavage-stage embryo. However, it remains uncertain whether this advantage extends to specialized subpopulations, such as women with thin endometrium (TE), who are characteristic of impaired endometrial receptivity. Thus, this study aims to evaluate the difference in the CLBR between cleavage-stage and blastocyst-stage embryos at fresh ET specifically in women with TE.MethodsA retrospective cohort comprising 1089 women from three centers, ranging from September 2017 to January 2022, was established. These women were diagnosed with TE (defined as endometrium thickness

Background: There is a definite shift in assisted reproductive centres from cleavage-stage embryo transfer (ET) toblastocyst transfer that is attributed to improvements in laboratory environments and advances in the development ofembryo culture media. The aim of the study was to investigate the reproductive outcomes of thawed cleavage-stage ET versusblastocysts derived from an extended culture of these embryos.Materials and Methods: This open-label, randomised, parallel group clinical trial study enrolled 182 women aged ≤37years who underwent frozen-thawed ET from November 2015 to June 2020 at Royan Institute Research Centre, Tehran,Iran. The women were randomly assigned to either the thawed cleavage ET group (n=110) or the post-thaw extended cultureblastocysts group (n=72). The primary outcome measure was the clinical pregnancy rate. Secondary outcome measureswere implantation rate, live birth rate (LBR), and miscarriage rate. A P

Background: There is a definite shift in assisted reproductive centres from cleavage-stage embryo transfer (ET) to blastocyst transfer that is attributed to improvements in laboratory environments and advances in the development of embryo culture media. The aim of the study was to investigate the reproductive outcomes of thawed cleavage-stage ET versus blastocysts derived from an extended culture of these embryos. Materials and Methods: This open-label, randomised, parallel group clinical trial study enrolled 182 women aged ≤37 years who underwent frozen-thawed ET from November 2015 to June 2020 at Royan Institute Research Centre, Tehran, Iran. The women were randomly assigned to either the thawed cleavage ET group (n=110) or the post-thaw extended culture blastocysts group (n=72). The primary outcome measure was the clinical pregnancy rate. Secondary outcome measures were implantation rate, live birth rate (LBR), and miscarriage rate. A P<0.05 indicated statistical significance. Results: There were no significant differences between the two groups in terms of demographic characteristics. Both the mean numbers of embryos transferred and good quality embryos transferred were significantly lower in the postthaw extended culture blastocysts group compared to thawed cleavage-stage ET cycles. However, the post-thaw extended culture blastocysts group had higher clinical pregnancy (56.94 vs. 40.91%, P=0.034), implantation (34.43 vs. 19.84%, P=0.001) and live birth (49.3 vs. 33.63%, P=0.036) rates compared to the thawed cleavage-stage ET group. Miscarriage and multiple gestations rates were comparable between the groups. Conclusion: These results allow us to take a position in favour of post-thaw extended culture blastocysts; thus, it is important to improve the post-thawing extended culture technique (registration number: NCT02681029). [ABSTRACT FROM AUTHOR]

Summary Embryo transfer is a crucial step in IVF cycle, with increasing trend during the last decade of transferring a single embryo, preferably at the blastocyst stage. Despite increasing evidence supporting Day 5 blastocyst-stage transfer, the optimal day of embryo transfer remains controversial. The crucial questions are therefore, whether the mechanisms responsible to embryos arrest are embryo aneuploidy or others, and whether those embryos arrested in-vitro between the cleavage to the blastocyst stage would survive in-vivo if transferred on the cleavage-stage. We therefore aim to explore whether aneuploidy can directly contribute to embryo development to the blastocyst stage. Thirty Day-5 embryos, that their Day-3 blastomere biopsy revealed a single-gene defect, were donated by 10 couples undergoing preimplantation genetic testing treatment at our center. Affected high quality Day-3 embryos were cultured to Day-5, and were classified to those that developed to the blastocyst-stage and those that were arrested. Each embryo underwent whole genome amplification. Eighteen (60%) embryos were arrested, did not develop to the blastocyst stage and 12 (40%) have developed to the blastocyst stage. Nineteen embryos (63.3%) were found to be euploid. Of them, 12 (66.6%) were arrested embryos and 7 (58.3%) were those that developed to the blastocyst-stage. These figures were not statistically different (p = 0.644). Our observation demonstrated that the mechanism responsible to embryos arrest in vitro is not embryo aneuploidy, but rather other, such as culture conditions. If further studies will confirm that Day-5 blastocyst transfer might cause losses of embryos that would have been survived in vivo, cleavage-stage embryo transfer would be the preferred timing. This might reduce the cycle cancellations due to failure of embryo to develop to the blastocyst stage and will provide the best cumulative live birth-rate per started cycle.

Abstract Background Preimplantation genetic testing (PGT) is offered to a wide range of structural and numerical chromosomal imbalances, with PGT- polymerase chain reaction (PCR), as the method of choice for amplifying the small DNA content achieved from the blastomere biopsy or trophectoderm (TE) biopsy, that might have a detrimental impact on embryonic implantation potential. Since human embryos cultured until Day-5–6 were noticed to expel cell debris/ fragments within the zona pellucida, we aimed to examine whether these cell debris/ fragments might be used for PGT, as an alternative to embryo biopsy. Methods Blastocysts, which their Day-3 blastomere biopsy revealed an affected embryo with single-gene defect, and following hatching leaved cell debris/fragments within the zona pellucida were analyzed. Each blastocyst and its corresponding cell debris/fragments were separated and underwent the same molecular analysis, based on multiplex PCR programs designed for haplotyping using informative microsatellites markers. The main outcome measure was the intra-embryo congruity of Day-3 blastomere biopsy and its corresponding blastocyst and cell debris/fragments. Results Fourteen affected embryos from 9 women were included. Only 8/14 (57.2%) of embryos demonstrated congruent molecular genetic results between Day-3 embryo and its corresponding blastocyst and cell debris/fragments. In additional 6/14 (42.8%) embryos, molecular results of the Day-3 embryos and their corresponding blastocysts were congruent, while the cell debris/fragments yielded no molecular diagnoses (incomplete diagnoses). Conclusions It might be therefore concluded, that in PGT cycles, examining the cell debris/fragments on Day-4, instead of Day-3 blastomere or Day-5 TE biopsies, is feasible and might avoid embryo biopsy with its consequent detrimental effect on embryos’ implantation potential. Whenever the latter results in incomplete diagnosis, TE biopsy should be carried out on Day-5 for final genetic results. Further large well-designed studies are required to validate the aforementioned PGT platform.

Summary: Embryo transfer is a crucial step in IVF cycle, with increasing trend during the last decade of transferring a single embryo, preferably at the blastocyst stage. Despite increasing evidence supporting Day 5 blastocyst-stage transfer, the optimal day of embryo transfer remains controversial. The crucial questions are therefore, whether the mechanisms responsible to embryos arrest are embryo aneuploidy or others, and whether those embryos arrested in-vitro between the cleavage to the blastocyst stage would survive in-vivo if transferred on the cleavage-stage. We therefore aim to explore whether aneuploidy can directly contribute to embryo development to the blastocyst stage. Thirty Day-5 embryos, that their Day-3 blastomere biopsy revealed a single-gene defect, were donated by 10 couples undergoing preimplantation genetic testing treatment at our center. Affected high quality Day-3 embryos were cultured to Day-5, and were classified to those that developed to the blastocyst-stage and those that were arrested. Each embryo underwent whole genome amplification. Eighteen (60%) embryos were arrested, did not develop to the blastocyst stage and 12 (40%) have developed to the blastocyst stage. Nineteen embryos (63.3%) were found to be euploid. Of them, 12 (66.6%) were arrested embryos and 7 (58.3%) were those that developed to the blastocyst-stage. These figures were not statistically different (p = 0.644). Our observation demonstrated that the mechanism responsible to embryos arrest in vitro is not embryo aneuploidy, but rather other, such as culture conditions. If further studies will confirm that Day-5 blastocyst transfer might cause losses of embryos that would have been survived in vivo, cleavage-stage embryo transfer would be the preferred timing. This might reduce the cycle cancellations due to failure of embryo to develop to the blastocyst stage and will provide the best cumulative live birth-rate per started cycle. [ABSTRACT FROM AUTHOR]

Background: Preimplantation genetic testing (PGT) is offered to a wide range of structural and numerical chromosomal imbalances, with PGT- polymerase chain reaction (PCR), as the method of choice for amplifying the small DNA content achieved from the blastomere biopsy or trophectoderm (TE) biopsy, that might have a detrimental impact on embryonic implantation potential. Since human embryos cultured until Day-5–6 were noticed to expel cell debris/ fragments within the zona pellucida, we aimed to examine whether these cell debris/ fragments might be used for PGT, as an alternative to embryo biopsy. Methods: Blastocysts, which their Day-3 blastomere biopsy revealed an affected embryo with single-gene defect, and following hatching leaved cell debris/fragments within the zona pellucida were analyzed. Each blastocyst and its corresponding cell debris/fragments were separated and underwent the same molecular analysis, based on multiplex PCR programs designed for haplotyping using informative microsatellites markers. The main outcome measure was the intra-embryo congruity of Day-3 blastomere biopsy and its corresponding blastocyst and cell debris/fragments. Results: Fourteen affected embryos from 9 women were included. Only 8/14 (57.2%) of embryos demonstrated congruent molecular genetic results between Day-3 embryo and its corresponding blastocyst and cell debris/fragments. In additional 6/14 (42.8%) embryos, molecular results of the Day-3 embryos and their corresponding blastocysts were congruent, while the cell debris/fragments yielded no molecular diagnoses (incomplete diagnoses). Conclusions: It might be therefore concluded, that in PGT cycles, examining the cell debris/fragments on Day-4, instead of Day-3 blastomere or Day-5 TE biopsies, is feasible and might avoid embryo biopsy with its consequent detrimental effect on embryos' implantation potential. Whenever the latter results in incomplete diagnosis, TE biopsy should be carried out on Day-5 for final genetic results. Further large well-designed studies are required to validate the aforementioned PGT platform. [ABSTRACT FROM AUTHOR]

Objective: To evaluate the effect of embryo stage at transfer on placental histopathology and perinatal outcome in singleton live births resulting from fresh embryo transfers (ETs).Design: Retrospective cohort study.Setting: Not applicable.Patient(s): The study population included all live births after fresh ETs during the period from 2009 to 2017.Intervention(s): None.Main Outcome Measure(s): Primary outcomes included anatomic, inflammatory, vascular malperfusion, and villous maturation placental features. Secondary outcomes included fetal, maternal, and perinatal complications.Result(s): A total of 677 live births were included in the final analysis and were allocated to the cleavage-stage (n = 252) and blastocyst (n = 425) ET groups. After the adjustment for confounding factors, the blastocyst group was found to be associated with a higher incidence of circummarginate membranes insertion (odds ratio [OR] 1.9, 95% confidence interval [CI] 1.2-3.4), delayed villous maturation (OR 8.5, 95% CI 1.2-69.3), chorangiosis (OR 2.0, 95% CI 1.2-3.8), parenchymal calcifications (OR 10.6, 95% CI 1.4-80.2), and intrapartum nonreassuring fetal heart rate tracing (OR 2.4, 95% CI 1.3-4.5). Compared with cleavage-stage ETs, live births resulting from the blastocysts were associated with a lower incidence of velamentous cord insertion (OR 0.5, 95% CI 0.3-0.9), retroplacental hematoma (OR 0.3, 95% CI 0.1-0.8), subchorionic thrombi (OR 0.3, 95% CI 0.1-0.8), and avascular villi (OR 0.2, 95% CI 0.03-0.7).Conclusion(s): Live births resulting from fresh cleavage-stage and blastocyst ETs have different placental histopathology features, with a higher rate of intrapartum nonreassuring fetal heart rate tracing in the blastocyst group. [ABSTRACT FROM AUTHOR]

Purpose: The present post hoc analysis aims to study the neonatal data of singletons born from three randomized controlled trials (RCTs) which compared the outcome of day 3 and day 5 transfers. Methods: Our analysis included 208 liveborn singletons from three existing RCTs (publication dates 2004, 2005, and 2006), 93 children from cleavage-stage transfers and 115 from blastocyst-stage transfers. Vanishing twins were excluded from the analysis. Singleton birthweight was the primary outcome measure. Gestational age and gender of the newborn were accounted for in the multiple regression analysis, along with other confounding factors, such as maternal age, BMI, parity, and smoking behavior. Results: There was no significant difference in gestational age (median, interquartile range) between cleavage-stage transfer (275 days; 267–281) and blastocyst-stage transfer (277 days; 270–281; p = 0.22). Singleton birthweight (median, interquartile range) was not significantly different between cleavage-stage transfer (3330 g; 3020–3610) and blastocyst-stage transfer (3236 g; 2930–3630; p = 0.40), even following multivariable regression analysis to control for potential maternal and newborn confounders. Conclusion: The gestational age and birthweight were not significantly different after cleavage-stage and blastocyst-stage transfers. One limitation to be recognized is the age of the data, with original data collection dates from 2001 to 2004. Additionally, the RCTs used for the present analysis have a fairly young age restriction. [ABSTRACT FROM AUTHOR]

Background: The incidence of monozygotic twins (MZT) after ART appears to be higher than the incidence after spontaneous conceptions contradicting the aim of ART to avoid multiple pregnancies because of the associated risks.Objective and Rationale: The aim was to study the frequency of MZT after IVF and ICSI and how it is influenced by the day of embryo transfer, maternal age, zona pellucida manipulation, controlled ovarian stimulation, stimulation protocol, culture media and embryo quality.Search Methods: Original studies and reviews were identified by searching the PubMed, Embase and Cochrane databases up to March 2017. The inclusion criterion was publications focusing on the five study questions related to MZT in our study. The exclusion criteria were articles that did not include blastocyst transfer, were on non-humans, were not published in peer-reviewed journals, and were based only on case studies. All of the articles were categorized according to the Oxford Centre for Evidence-based Medicine's 'Levels of Evidence', and quality and risk of bias assessment was performed with 'The Cochrane Collaboration's Risk of Bias Tools'. A meta-analysis was performed to study the impact of the day of embryo transfer on the MZT rate.Outcomes: The literature search resulted in a total of 42 articles, including 38 original studies, for analysis. The included original studies reported a MZT rate with blastocyst transfer from zero to 13.2%. Our meta-analysis found a higher frequency of MZT after blastocyst transfer compared with cleavage-stage embryos transfer: odds ratio = 2.18, 95% CI: 1.93-2.48 (fixed effect meta-analysis). A younger maternal age may increase the MZT rate, and recent studies regarding the use of zona pellucida manipulating techniques have disagreed with the previous suspicion of a higher MZT rate after the use of these methods. The extended culture to-blastocyst stage is a potential risk factor for MZT, but it is uncertain whether this phenomenon is due to the extended time, culture media or greater likelihood of younger oocytes to reach the blastocyst stage. An increased frequency of MZT following the GnRH-agonist suppression protocol has been suggested, as well as a decreased frequency of MZT with high gonadotrophin doses, which could reflect an age-related effect. Only limited literature has focused on the role of embryo morphology in the MZT rate, therefore, this issue remains unresolved.Wider Implications: We found blastocyst transfer to be a risk factor for MZT. Hence, the results of this meta-analysis may weaken the previously proposed view that greater experience with blastocyst transfer and improved culture media could decrease the high rate of MZT after blastocyst transfer. To minimize the rate of MZT and the associated complications, the mechanisms underlying blastocyst transfer and MZT pregnancy must be elucidated. [ABSTRACT FROM AUTHOR]

Study Question: Do cumulative live birth rates differ between single cleavage-stage Day 3 transfer and single blastocyst-stage Day 5 transfer?Summary Answer: Cumulative live birth rates after Day 3 and 5 transfers were similar in young patients when the vitrified embryo transfers were also taken into account.What Is Known Already: Previous evidence has shown that the probability of live birth following IVF with a fresh embryo transfer is significantly higher after blastocyst-stage Day 5 transfer. However, because the introduction of vitrification has enhanced the survival of cryopreserved embryos and improved pregnancy rates, the optimal outcome measure for this comparison should now be cumulative live birth rates, as these include the eventual contribution of vitrified-warmed embryos.Study Design, Size, Duration: Our retrospective study included first IVF/ICSI cycles performed between January 2010 and December 2013 at a tertiary care centre.Participants/materials, Setting, Methods: All patients were scheduled for fresh single embryo transfer, either on Day 3 (n = 377) or on Day 5 (n = 623). Both IVF and ICSI cycles were included and the sperm used were either fresh or frozen partner ejaculates, or frozen donor ejaculates. The primary outcome was cumulative live birth (after 24 weeks) rate per started cycle, including the eventual contribution of vitrification until the birth of a first child.Main Results and the Role Of Chance: Live birth rates per started cycle were significantly lower after transferring the fresh single cleavage-stage embryo, compared to a blastocyst (31.3% and 37.8%, respectively, P = 0.041). Furthermore, the number of embryo transfers necessary until the first live birth was significantly lower for blastocyst-stage embryos (P < 0.001). However, the cumulative live birth rates were 52.6% for cleavage-stage and 52.5% for blastocyst-stage transfers (P = 0.989).Limitations, Reasons For Caution: The extrapolation of the results is limited by the retrospective nature of the study. Furthermore, the analysis was restricted to patients under 36 years of age undergoing their first treatment cycle.Wider Implications Of the Findings: These results deserve further clinical consideration in terms of time and cost efficiency. A subsequent analysis of the neonatal outcomes is necessary to confirm the safety of treatment cycles using extended culture.Study Funding/competing Interests: No external funding was received and there are no conflicts of interest to declare. [ABSTRACT FROM AUTHOR]

Purpose: The aim of this study was to explore whether transferring two grades I cleavage-stage embryo was suitable for the patients in the first fresh transfer. Methods: This study included 202 single grades I cleavage-stage, 229 single grades III cleavage-stage, 743 single excellent blastocyst, 522 double grades I cleavage-stage, and 596 double grades III cleavage-stage embryo transfers. Main clinical outcomes: clinical pregnancy and twin-pregnancy rate. Results: Among single excellent blastocyst, single grades I and single grades III group, the clinical pregnancy rate was significantly higher in single excellent blastocyst group than single grades I and grades III group (67.16% versus 42.08% versus 23.97%;p < 0.001). When transferred double grades I cleavage-stage embryos, the clinical pregnancy rate reached 68.20% which was no significant difference compared with the single excellent blastocyst group (67.16%). However, the twin-pregnancy rate was significantly higher in double grades I group than double grades III and single excellent blastocyst group (43.26% versus 26.70% versus 0.60%;p < 0.001). Conclusions: Because of higher twin-pregnancy incidence rate, transferring two grades I cleavage-stage embryo might not be a good protocol. Extended culture to blastocyst-stage could be considered for the patient with only two grades I cleavage-stage embryos. [ABSTRACT FROM AUTHOR]

Assessment of the development ability of oocytes following vitrification and thawing is an important step for the optimization of "in vitro" fertilization techniques. The classic "in vitro" fertilization of vitrified-thawed oocytes in mice is often inefficient because of incomplete capacitation of spermatozoa in the absence of surrounding cumulus cells. The aim of this study was to compare the fertilization and embryo development efficiency of vitrified-thawed oocytes assessed by two modem "in vitro" fertilization methods: Intracytoplasmic sperm injection (ICSI) and laser assisted IVF (ZD-IVF) and to overcome the incomplete capacitation of spermatozoa and the zona pellucida hardening following vitrification. A total of 110 oocytes, obtained from superovulated B6D2F1 females were vitrified using the OPS method and subsequently warmed on a later date. Surviving oocytes (n=96, 87.3%) were randomly allocated for fertilization by ICSI (n=56, group 1) or laser assisted IVF (n=60, group 2) using fresh spermatozoa from CD1 stud males. Fertilization was achieved in 37 (66%) of the oocytes from group 1 and 38 (63%) from group 2. The developmental rate of 4 cells stage embryos, morulae and blastocysts (36±60%, 35±58.5%, 19±53%) in the first group was similar comparing with the second group (34±60.7%, 32±57.1%, 28±50%). We conclude that the fertilization and embryo development efficiency of vitrified-thawed oocytes can be enhanced significantly using ICSI or Laser Assisted IVF. These data also suggest that by using one of these methods we can overcome the incomplete capacitation of spermatozoa and the zona pellucida hardening following vitrification. [ABSTRACT FROM AUTHOR]

Objective: To compare the incidence of monozygotic twinning between cleavage-stage and blastocyst-stage embryo transfer in a large cohort of patients undergoing single embryo transfer. Design: Retrospective study. Setting: Dutch-speaking Free University of Brussels. Patient(s): This study covered the period between July 2003 and December 2005. 1,951fresh IVF/ICSI cycles in which single embryo transfer was performed were retrospectively reviewed. Only the first cycle of each patient was included. Intervention(s): Five hundred eighty seven (n = 587) cycles that resulted in clinical pregnancies were identified; 308 after single day-3 embryo transfer and 271 after single blastocyst transfer. Main Outcome Measure(s): The incidence of monozygotic twinning. Result(s): Overall, 13 cases (2.2%) of monozygotic twinning were observed, 2.6% in the cleavage-stage group (n = 8/308) and 1.8% in the blastocyst group (n = 5/271). No statistically significant differences were observed in the probability of monozygotic twinning between the Cleavage-stage and the Blastocyst group (difference: +0.8%; 95% CI, –1.97 to +3.41). All of these pregnancies resulted in the delivery of 24 healthy babies. The crude odds ratio for the incidence of monozygotic twinning after day-5 embryo transfer was calculated to be 0.71 (95% CI, 0.23–2.18). Conclusion(s): To investigate the potential association between the day of embryo transfer (day 3 or 5) and the incidence of monozygotic twinning, the clinical pregnancies analyzed should have been established after single embryo transfer. The current study represents the first methodologically appropriate study attempting to investigate the above research question. Our findings support that opting for blastocyst transfer does not increase the probability for monozygotic twins. [Copyright &y& Elsevier]

A total of 1612 expression sequence tags derived from Ciona intestinalis cleavage-stage embryos were examined to explore detailed gene expression profiles. The 3′ sequences indicate that the 1612 clones can be categorized into 1066 independent clusters. DDBJ database search suggested that 496 of them showed significant matches to reported proteins with distinct functions. Among them 69 are associated with cell–cell communications and 41 with transcription factors. In situ hybridization of all 1066 clusters showed that 84 clusters exhibited blastomere-specific pattern of expression, and many of these genes seem to encode for novel proteins. One of the interesting findings is that most of them were expressed in the precursor cells of multiple tissues. Among them 28 genes were expressed in the marginal zone of the 32-cell embryo. The blastomeres in this region are thought to receive an inductive signal from the vegetal blastomeres. Many of the blastomere-specific genes did not show similarity to known proteins. The present analysis therefore provides new information for further analyses on the cell fate specification in the Ciona embryo. [Copyright &y& Elsevier]