Biomedical subjects

V Gindilis

Publications and source records attributed to V Gindilis.

7 recordsLinked to original sources

PubMed1999

Construction and sequence analysis of subtraction complementary DNA libraries from human preimplantation embryos.

PURPOSE: Because stage-specific genetic expression in human preimplantation development is not sufficiently studied, we have undertaken the construction of a subtraction complementary DNA (cDNA) library enriched for transcripts specific for human blastocysts. METHODS: For this purpose individual pools of cDNAs synthesized from four hatched blastocysts and three cleaving 8- to 10-cell embryos were exposed to suppression subtractive hybridization to minimize the presence of transcripts of housekeeping genes and other genes of maternal origin known to be expressed earlier in preimplantation development. Random clones of this library were sequenced and analyzed using the BLAST algorithm. RESULTS: The resulting subtraction library had a complexity of 3 x 10(5) and an average size of inserts of about 0.8 kb. Sequencing of random library clones revealed the following human genes: CD9 antigen, fatty acid binding protein, ferritin heavy chain, amyloid precursor, MAP kinase messenger RNAs, DNA clone 127H14, messenger RNA for diacylglycerol kinase, a sequence homologous to C1 inhibitor, messenger RNA for the KIAA0145 gene, and others. CONCLUSIONS: The presence of these genes in human preimplantation development suggests expression specific to the blastocyst stage.

Algorithms↗

PubMed1998

Sequence analysis of libraries from individual human blastocysts.

PURPOSE: It has recently become possible to construct cDNA libraries from individual human blastocysts to investigate the expression of embryonic genes in human preimplantation development. We have previously reported the expression of beta-actin, CD-59, and homeobox OCT-3 and identified almost-complete homology of sequences to human histone 3.1 and human ribosome protein S25. In the present paper, our further sequencing analysis of cDNA libraries from single human blastocysts is described. METHODS: cDNA libraries were constructed from 13 blastocysts. Sequence analysis was performed in 120 clones from one of these cDNA libraries with fragments of 50 to 1000 bp. Their sequence identity was analyzed using the expressed sequence tag (EST) database. RESULTS: The presence of two housekeeping genes, hexokinase I and serine/threonine phosphorylase, and four other ESTs was demonstrated, the identity of which, with particular gene expression in preimplantation development, has not yet been established. CONCLUSIONS: The data demonstrate the usefulness of constructing cDNA libraries from individual human blastocysts and their values in the analysis of genetic expression in human preimplantation development.

Blastocyst↗

PubMed1998

Evolutionary classification of homeodomains.

PURPOSE AND METHODS: We performed multiple comparisons between available amino acid (aa) sequences of homeodomain(HOM)-containing proteins from a wide spectrum of animals to create an evolutionary classification of the proteins. RESULTS: Based on results of statistical and special computational analyses of over 500 homeodomain aa sequences (HOMs) a novel system of concepts describing complex structural correlations between homologous proteins is proposed. This system includes such notions as differentiated isofunctionality of aa, chemotype, stereotype, local functional motifs, gradual conservativeness of aa positions, and group-specific domain patterns, as well as major categories of the evolutionary classification of HOMs (Division, Type, Branch, Class, Family, Series, Variety, Sort). Using this approach, a complete structural systematics of HOMs belonging to proteomes of eukaryotic animals is proposed. CONCLUSIONS: The proposed structural classification of HOMs is in full agreement with the bulk of experimental data revealing complex functional similarities and differences among HOMs in terms of their expression patterns in developing embryos. It turn, this classification can provide answers regarding homology among homeodomains when experimental data are conflicting.

Amino Acid Sequence↗

PubMed1998

Isolation of cDNA libraries from individual human preimplantation embryos.

Although available data from the mouse model suggest that morphogenesis during preimplantation development is dependent on the expression of embryonic genes, convincing data for human preimplantation embryos are missing. To investigate the expression of genes involved in human preimplantation development we constructed cDNA libraries from human individual blastocysts and screened them for the expression of beta-actin, CD59, homeoboxes OCT-3 and HOXA4, and HLA-G and hMLH-1 genes. Beta-actin, CD59, and OCT-3 were detected by reverse transcription-polymerase chain reaction (RT-PCR), while HOXA4, HLA-G and hMLH-1 were undetected. Sequencing of 48 random clones from two libraries revealed a different identity to the known genes including 99% identity to human histone 3.1 and human ribosome protein S25 complete cDNA. These data demonstrate the feasibility of constructing cDNA libraries from individual human preimplantation embryos and their potential usefulness in the assessment of the relevance of specific gene expression in the failures of preimplantation development.

Blastocyst↗

PubMed1996

Expression of homebox-containing genes in human preimplantation development and in embryos with chromosomal aneuploidies.

PURPOSE: The purpose of the study was to investigate homeobox gene expression in human oocytes and preembryos and in postimplantation embryos with impaired embryonic development determined by chromosomal abnormalities. METHODS: Reverse transcriptase-polymerase chain reaction (RT-PCR) with intron spanning primer sets for Homeobox gene sequences was used. RESULTS: The homeobox genes HoxA4, HoxA7, HoxB4, and HoxB5 were present in human oocytes and cleaving normal and triploid embryos. The expression pattern was different between chromosomally abnormal and normal first-trimester embryos. Of four homeobox transcripts (HoxA7, HoxB4+ ++, HoxB5, and HoxC6) that are expressed in diploid embryos, only HoxA7, HoxB4 and HoxC6 were present in a trisomy 7 embryo, and only HoxB4 and HoxB 5 in triploid embryos and an embryo with trisomy 9. Cloning experiments revealed differences in the number of homeobox clones obtained from trisomy 7 and control embryos. CONCLUSIONS: The transcripts of homeobox genes, HoxA4, HoxA7, HoxB4, and HoxB5, were present in oocytes and cleaving embryos. The pattern of expression of homeobox genes in cultured fibroblasts derived from spontaneously aborted embryos with aneuploidies was different from that in control diploid cells.

Abortion, Spontaneous↗

PubMed1995

Homeobox gene expression in human oocytes and preembryos.

Homeobox gene expression in human preimplantation development has not been established. We used reverse transcriptase-polymerase chain reaction (RT-PCR) with intron spanning primer sets to investigate the presence of mRNA of homeobox genes in human oocytes and preembryos. RT-PCR products obtained from normal and unfertilized oocytes, from cleaving normal and triploid embryos, and from morulae and blastocysts were cloned, sequenced, and analyzed for the presence of homeobox sequences. The presence of mRNA of homeoboxes HoxA4 and HoxA7 was demonstrated; HoxA4 was present in normal and unfertilized oocytes and also in a 4-cell embryo. HoxA7 was present in normal oocytes and cleaving triploid embryos.

Amino Acid Sequence↗

PubMed1994

Review: borders, patterns, and distinctive families of homeodomains.

PURPOSE: Homeotic proteins function as transcription factors in early embryogenesis of many organisms. To date, hundreds of distinctive homeoproteins have been identified, including 84 human homeodomains. However further progress in understanding functional relationships between particular homeoproteins and other embryonic regulators requires a comprehensive structural classification of these proteins. RESULTS: The most probable borders and conservative amino acid positions inside the homeodomain region have been established using a statistical analysis of variabilities of amino acid occurrences at various positions outside and inside the domain. A new format for a homeodomain sequence presentation and regular amino acid patterns which are strongly representative of distinctive homeodomain groups are proposed. Using the established patterns, 33 families of closely related homeodomains have been distinguished and classified. The total list of 297 homeodomain amino acid sequences is presented in the Appendix. CONCLUSION: The structural classification of homeodomains has been proposed. It can be useful for both the identification (or prediction) of new homeotic genes/proteins and the recognition of possible PCR-induced sequence errors. This systematics will also have an impact on understanding functional relationships among homeotic proteins and other genetic regulators of developmental processes.

Amino Acid Sequence↗