User:Z3414515

From Embryology

Lab Attendance

Lab 1 --Z3414515 (talk) 12:46, 6 August 2014 (EST)

Lab 2 --Z3414515 (talk) 11:14, 13 August 2014 (EST)

Lab 3 --Z3414515 (talk) 11:07, 20 August 2014 (EST)

Practice

http://www.ncbi.nlm.nih.gov/pubmed

PubMed

Pmid4118885

<pubmed>4118885</pubmed>

My Type in a Group

Teamworker

A Teamworker is the oil between the cogs that keeps the machine that is the team running smoothly. They are good listeners and diplomats, talented at smoothing over conflicts and helping parties understand one other without becoming confrontational. Since the role can be a low-profile one, the beneficial effect of a Teamworker can go unnoticed and unappreciated until they are absent, when the team begins to argue, and small but important things cease to happen. Because of an unwillingness to take sides, a Teamworker may not be able to take decisive action when it is needed.

Lecture Reviews

Lecture 1

Course introduction for embryology as well as the history of embryologists and how the diagrams of embryo changed through time as more advance technology was available. Guidelines to the course was mentioned as well as the assessments and type of work expected for this course.

Lecture 2

In the fertilization lecture the most interesting concept for me was the polar bodies and the sry gene. Every other concepts such as gametes, mitosis, meiosis and fertilization was familiar. Polar bodies and the sry gene was a completely new idea for me. Meiosis 1 releases first polar body and meiosis 2 releases the second polar body. Sometimes meiosis 1 releases first and third polar bodies.

Individual Assessments

Lab 1

Reference: Pmid25089626

<pubmed>25089626</pubmed>

Method summary

Microarray Analysis

Raw data on Affymetrix GeneChip HGU133 were obtained from the ArrayExpress for human preimplantation embryos. The invariant set normalisation method was used and via using the Li-Wong method, the expression values were extracted from PM-values. The arrays were normalised independently and Li-Wong method was applied to all normalised arrays to get a summary of the expression measurements. Using Bayesian approach, differential expression between the consecutive development stages was analysed.

Embryo Collection

FVB/N mice were kept for 12 hours under light/dark cycle and were fed regularly. A Pregnant Mare’s Serum (5 IU) was injected into a 4-7 weeks old female. After 44 hours a human chorionic gonadotropin (5 IU) injection was given. The females then mated with the FVB/N strain studs (males). 19-21 hours later the females were sacrificed and the oviducts were collected. Oocytes were collected. The embryos were then cultured in KSOM medium.

Gene expression analysis

Extraction of RNA from mouse unfertilised oocytes using Arcturus PicoPure RNA isolation kit was done. Agilent Bioanalyser was used to measure the RNA quality and concentration. One embryo yielded 128 pg of total RNA on average. For each final protocol, three biological replicas of all the stages were collected.

TaqMan Array Cards analysis

RQ Manager version 1.2.2 (Applied Biosystems) were used to analyse Ct values. Hprt1 and Psmb6 were the endogenous controls which were used for normalisation.

Expression analysis from public sequencing dataset

Gene Expression Omnibus database was used to obtain the normalised RPKM values for human and mouse pre-implantation stages. The p-values were calculated for the pairs i.e. oocytes and 4-cell blastomeres and etc. The p-values below 0.05 were significant. In human and mouse, the average values for each stage between embryos in the same biological stages were calculated.

Result summary

Analysis of two independent human pre-implantation microarray datasets were done in order to define the genes with consistent gene expression profiles between embryo stages. The probes which had significant changes in both datasets were considered for further analysis. Probes in the “Up-down” cluster were up regulated whereas the probes in “Down” cluster were down regulated. Genes were selected from each cluster “Up”, Up-down” and “Down” for analysis of expression profile of mouse pre-implantation embryo by qPCR. A gene was included if its ortholog was found in any of the following samples in MGI: oocyte, unfertilized oocyte, fertilized oocyte, 2-cell embryo, 4-cell embryo, 8-cell embryo, 16-cell embryo, blastocyst. In the mouse, 55 genes with orthologs were selected for gene expression profiling. Also expression patterns of the selected genes in the mouse were studied. The maternal gene expression profile was seen to be shared in more than half of the mouse orthologs for genes “Up” and “Up-down” clusters. All the PRAME and most SSX, MAGEA and GAGE family members in human microarray were of “up-down” cluster. However, in the pre-implantation human embryo, the selected families’ genes had dynamic expression profiles.

Reference: Pmid25071849

<pubmed>25071849</pubmed>

Method summary

This study was performed in Assisted Reproduction of Wuhan Union Hospital from January 2012 to December 2012. A total of 1891 cycles were used which contained 1150 fresh embryo transfers and 741 frozen-thawed embryo transfers. Cleavage-stage or blastocyst-stage was composed in 1150 women. Also 741 women were divided into cleavage-stage or cleavage-stage extended blastocyst culture or blastocyst-stage transfer. A GnRH agonist protocol was used in all the cycles. An injection of 10000 units of HCG was given to two or more follicles when they reached 18mm in diameter and then 34-36 hours later an ovum pick up was performed. After OPU, 4-6 hours later in vitro fertilisation was performed. The assessment for the embryo was based on the rate of development and morphology. All the good embryos were cryopreserved through vitrification. The number of implantations was observed as the number of sacs. Using the SPSS software, all the statistical calculations were performed.

Result summary

Clinical pregnancy rates for patients less than 35 years of age:

  • Fresh cleavage-stage embryo transfers: 52.7%
  • Fresh blastocyst transfers: 35.88%
  • Frozen-thawed cleavage-stage embryo transfers: 35.29%
  • Post thaw cleavage-stage extended blastocyst culture transfers: 47.75%
  • Frozen-thawed blastocyst transfers: 59.8%

Clinical pregnancy rates for patients more than 35 years of age:

  • Fresh cleavage-stage embryo transfers: 41.24%
  • Fresh blastocyst transfers: 26.92%
  • Frozen-thawed cleavage-stage embryo transfers: 11.32%
  • Post thaw cleavage-stage extended blastocyst culture transfers: 46.15%
  • Frozen-thawed blastocyst transfers: 55.8%

Lab 2

E18.5 developing kidney expressing Pygo1 and Pygo2.jpg

E18.5 developing kidney expressing Pygo1 and Pygo2 Expression patterns of Pygo1 and Pygo2 proteins in the cortex of E18.5 kidney was determined using immunofluorescence. The location of both Pygo1 and Pygo2 were in the nucleus with the colour red. Both genes are expressed widely where in all the components of the developing kidney, a signal is detected. However their were high levels of stromal cell compartment(arrows). Original magnification x200

Student number: 3414515

Reference

<pubmed>17425782</pubmed>

© 2007 Schwab et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


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