Scientific References: Human Genetic Engineering
(including Cloning): Artificial Embryos, Oocytes, Sperms, Chromosomes and Genes

B. Artificial Spermatogonia, Spermatocytes, and Sperms

-- Hum Mol Genet. 1992 Dec;1(9):717-26

Molecular isolation and characterization of an expressed gene from the human Y chromosome. Zhang JS, Yang-Feng TL, Muller U, Mohandas TK, de Jong PJ, Lau YF. (USA)

Using a positional cloning approach, we have isolated an expressed gene from a flow-sorted Y chromosome cosmid library. Several cDNA clones were isolated from a human testis cDNA library constructed in lambda gt10. In addition, DNA-mediated gene transfer and restriction enzyme mapping experiments demonstrated that two functional transcriptional units are present within the Y-231 cosmid. The Y-231 (TSPY) gene is conserved in the male genome and expressed in the testis of the chimpanzee, suggesting that it may play an important role in the physiology of this organ in man and the great ape. [PMID: 1284595]

-- Genomics. 1991 Sep;11(1):108-14

Cloning and sequence analysis of a human Y-chromosome-derived, testicular cDNA, TSPY. Arnemann J, Jakubiczka S, Thuring S, Schmidtke J. (UK)

The human Y-specific gene TSPY (testis-specific protein Y-encoded) was originally defined by the genomic probe pJA36B2 (DYS14), which detects a poly(A)+ RNA transcript in human testis tissue. Using this probe we have now isolated the cDNA sequence pJA923 from a human testis cDNA library PCR analysis of genomic DNA from patients with specific primers confirmed the simultaneous presence of at least two independent loci on the proximal short arm of the Y chromosome. [PMID: 1765369]

-- Biochem Biophys Res Commun. 1999 May 27;259(1):60-6

Cloning and characterization of SOB1, a new testis-specific cDNA encoding a human sperm protein probably involved in oocyte recognition. Lefevre A, Duquenne C, Rousseau-Merck MF, Rogier E, Finaz C. (France)

A human sperm-oocyte binding protein, SOB1, was purified .... This is a new testis-specific cDNA. It is a single copy gene, well conserved among mammals and located on human chromosome 12 at band p13. [PMID: 10334916]

-- Mol Reprod Dev. 1993 Dec;36(4):407-18

Sequence, expression, and chromosomal assignment of a human sperm outer dense fiber gene. Gastmann O, Burfeind P, Gunther E, Hameister H, Szpirer C, Hoyer-Fender S. (Germany)

Outer dense fibers (ODFs) are located on the outside of the axoneme in the midpiece and principal piece of the mammalian sperm tail and may help to maintain the passive elastic structures and elastic recoil of the sperm tail. We have identified and describe here a human gene that is homologous to the Mst(3)CGP gene family of Drosophila melanogaster and encodes an ODF protein of 241 amino acids. The gene is expressed in testis but not in human spleen, kidney, or brain. The isolation of a human gene encoding a sperm tail protein may provide the ability to identify and investigate, on the molecular level, possible reasons for human male infertility that are dependent on flagellar disturbances. [PMID: 8305202]

-- Genomics. 1993 Sep;17(3):736-9

Evidence that the SRY protein is encoded by a single exon on the human Y chromosome. Behlke MA, Bogan JS, Beer-Romero P, Page DC. (USA)

To facilitate studies of the SRY gene, a 4741-bp portion of the sex-determining region of the human Y chromosome was sequenced and characterized. Within this CpG island lies the sequence CGCCCCGC, a potential binding site for the EGR-1/WT1 family of transcription factors, some of which appear to function in gonadal development. [PMID: 8244390]

-- Genomics. 1995 Oct 10;29(3):796-800

Expression analysis, genomic structure, and mapping to 7q31 of the human sperm adhesion molecule gene SPAM1. Jones MH, Davey PM, Aplin H, Affara NA. (UK)

Nucleotide sequence for 1919 bp of human DNA from a series of overlapping cDNA clones isolated from a testis cDNA library confirmed the sequence identity within a 1527-bp open reading frame ... Northern analysis of poly(A)+ mRNA from a range of 16 human tissues has demonstrated that expression of the gene as a single 2.4-kb transcript is strictly limited to the testis. [PMID: 8575780]

-- Arch Androl. 1999 Mar-Apr;42(2):71-84

Molecular cloning and characterization of a novel testis-specific nucleoporin-related gene. Wang LF, Zhu HD, Miao SY, Cao DF, Wu YW, Zong SD, Koide SS. (Beijing, PRC)

RSD-1 was used as a probe to screen a human testis lambda ZAPII cDNA expression library. Northern blot analysis of mRNAs prepared from various human tissues shows that BS-63 is transcribed in two forms: 6.0 and 8.5 kb. The 8.5-kb transcript was present in low amounts in several somatic tissues, whereas the 6.0-kb transcript is expressed only in testis. In situ hybridization analysis of human testis sections showed that BS-63 mRNA is expressed only in germ cells at all stages of spermatogenesis. Sertoli cells did not transcribe the gene. [PMID: 10101573]

-- Genomics. 1994 Jul 1;22(1):205-10

The identification of novel gene sequences of the human adult testis. Affara NA, Bentley E, Davey P, Pelmear A, Jones MH. (UK)

To facilitate the characterization of genetic expression in human adult testis, expressed sequence tag analysis of cDNAs from this tissue has been undertaken. In comparison to similar studies on human brain tissue and a hepatoma cell line, the findings indicate that the matches in the testis transcript population appear to be identifying a different spectrum of gene sequences. [PMID: 7959769]

-- Anim Genet. 2002 Jun;33(3):211-4

Characterization of the porcine sperm adhesion molecule gene SPAM1 - expression analysis, genomic structure, and chromosomal mapping. Day AE, Quilter CR, Sargent CA, Mileham AJ. (UK)

Sequence analysis of cDNA products, derived from adult porcine testis mRNA, gave overlapping nucleotide sequence correlating to 1952 bp of the sperm adhesion molecule 1 (SPAM1) gene. This sequence was shown to be homologous to SPAM1 genes known in other mammalian species. Fluorescence in situ hybridization (FISH), using a bacterial artificial chromosome (BAC) clone from the PigE BAC library, was used to map SPAM1 to chromosome 18 of the pig. This finding is consistent with comparative mapping experiments performed between pig and human chromosomes. [PMID: 12030925]

-- Proc Natl Acad Sci U S A. 1993 Nov 1;90(21):10071-5

Molecular cloning of the human and monkey sperm surface protein PH-20. Lin Y, Kimmel LH, Myles DG, Primakoff P. (USA)

Here we report the isolation of human and cynomolgus monkey PH-20 cDNAs as a key step toward testing the function of primate PH-20 and the contraceptive efficacy of PH-20 immunization in primates. Southern blots show that there is a single PH-20 gene in the human genome and Northern blots of human testis poly(A)+ RNA show a 2.4-kb message. Northern blots of tissues other than testis are negative for PH-20, indicating that human PH-20 is testis-specific. [PMID: 8234258]

-- Biochem Biophys Res Commun. 1992 Oct 15;188(1):265-71

Molecular cloning and chromosomal mapping of the human gene for the testis-specific catalytic subunit of calmodulin-dependent protein phosphatase (calcineurin A). Muramatsu T, Kincaid RL. (USA)

A cDNA for an alternatively spliced variant of the testis-specific catalytic subunit of calmodulin dependent protein phosphatase (CaM-PrP) was cloned from a human testis library. Analysis of Southern blots containing DNA from human-hamster somatic cell hybrids show that the gene is on human chromosome 8. [PMID: 1339277]

-- Gene. 1998 Aug 17;216(1):31-8

The human glucosamine-6-phosphate deaminase gene: cDNA cloning and expression, genomic organization and chromosomal localization. Shevchenko V, Hogben M, Ekong R, Parrington J, Lai FA. (UK)

When mammalian eggs are fertilized by sperm, a distinct series of calcium oscillations are generated which serve as the essential trigger for egg activation and early embryo development. We have isolated the corresponding human testis homologue of the hamster sperm 33 kDa cDNA. The genomic structure of the human glucosamine-6-phosphate deaminase has been mapped and the gene was localized by fluorescence in situ hybridization (FISH) to chromosome 5q31. [PMID: 9714720]

-- J Cell Biol. 1990 Dec;111(6 Pt 2):2939-49

cDNA cloning reveals the molecular structure of a sperm surface protein, PH-20, involved in sperm-egg adhesion and the wide distribution of its gene among mammals. Lathrop WF, Carmichael EP, Myles DG, Primakoff P. (USA)

Sperm binding to the egg zona pellucida in mammals is a cell-cell adhesion process that is generally species specific. Cross-species Southern blots reveal the presence of a homologue of the PH-20 gene in mouse, rat, hamster, rabbit, bovine, monkey, and human genomic DNA, showing the PH-20 gene is conserved among mammals. [PMID: 2269661]

-- Gene. 2000 Oct 3;256(1-2):253-60

Human allantoicase gene: cDNA cloning, genomic organization and chromosome localization. Vigetti D, Monetti C, Acquati F, Taramelli R, Bernardini G. (Italy)

We have recently cloned the first vertebrate allantoicase cDNA from the amphibian Xenopus laevis. From a human fetal spleen cDNA library and adult kidney EST clone, we have obtained a 1790 nucleotide long cDNA. Allantoicase cDNA is expressed in adult testis, prostate, kidney and fetal spleen. By comparison with available genomic sequences deposited in database, we have determined that the human allantoicase gene consists of five exons and spans 8kb. We have also mapped the gene in chromosome 2. [PMID: 11054555]

-- Genomics. 2001 Oct;77(3):163-70

Cloning and chromosomal localization of a gene encoding a novel serine/threonine kinase belonging to the subfamily of testis-specific kinases. Visconti PE, Hao Z, Purdon MA, Stein P, Balsara BR, Testa JR, Herr JC, Moss SB, Kopf GS. (USA)

[W]e isolated a PCR fragment encoding a novel member of the testis-specific serine/threonine kinases (STK) from mouse male mixed germ cell mRNA. Northern blot analysis revealed that this protein kinase is developmentally expressed in testicular germ cells and is not present in brain, ovary, kidney, liver, or early embryonic cells. We then cloned the human homologue of this protein kinase gene (STK22C) and found it to be expressed exclusively in the testis. Fluorescence in situ hybridization with both the human and mouse cDNA clones revealed syntenic localization on chromosomes 1p34-p35 and 4E1, respectively. [PMID: 11597141]

-- Yi Chuan Xue Bao. 2003 Jan;30(1):25-9

Molecular cloning for testis spermatogenesis cell apoptosis related gene TSARG1 and Mtsarg1 and expression analysis for Mtsarg1 gene. Fu JJ, Lu GX, Li LY, Liu G, Xing XW, Liu SF. (Laboratory of Human Reproductive Engineering, Central South University Xiangya Medical College, Changsha, China)

Spermatogenesis cell apoptosis is a very complex process, which needs many molecules to take part in the programmable death of cells in testis. It is very important to clone spermatogenesis cell apoptosis related genes and spermatogenesis genes in testis. Applying the bioinformatics and experiment technique, we have cloned human and mouse novel gene cDNA sequences--human testis and spermatogenesis cell apoptosis related gene 1 (TSARG1) and mouse testis and spermatogenesis cell apoptosis related gene 1(Mtsarg1) from human and mouse testis cDNA library respectively. [O]ur results suggested that Mtsarg1 and TSARG1 would be pay potential roles in spermatogenesis cell apoptosis or spermatogenesis. [PMID: 12812072]

-- FEBS Lett. 1998 Mar 6;424(1-2):73-8

Cloning, expression analysis and chromosomal localization of the human nuclear receptor gene GCNF. Agoulnik IY, Cho Y, Niederberger C, Kieback DG, Cooney AJ. (USA)

Germ cell nuclear factor (GCNF) is an orphan member of the nuclear receptor gene superfamily. We report the cloning of a cDNA encoding a new variant of human GCNF from human testis and its expression analysis. Chromosomal localization of the GCNF gene shows that the gene is located on chromosome 9. In situ hybridization analysis of GCNF expression in the testis shows that human GCNF is expressed exclusively in germ cells. [PMID: 9537518]

III. Artificial Chromosomes and Genes

-- Gene. 2003 Nov 27;320:165-76

Barnacle: an assembly algorithm for clone-based sequences of whole genomes. Choi V, Farach-Colton M. (USA)

We propose an assembly algorithm Barnacle for sequences generated by the clone-based approach. We illustrate our approach by assembling the human genome. The assembly of December 2001 freeze of the public working draft generated by Barnacle and the source code of Barnacle are available at ( [PMID: 14597400]

-- Mol Cell Biol. 2003 Nov;23(21):7689-97

Human artificial chromosomes with alpha satellite-based de novo centromeres show increased frequency of nondisjunction and anaphase lag. Rudd MK, Mays RW, Schwartz S, Willard HF (USA)

Human artificial chromosomes have been used to model requirements for human chromosome segregation and to explore the nature of sequences competent for centromere function. [PMID: 14560014]

-- Science. 2001 Oct 5;294(5540):109-15; Comment in: Science. 2001 Oct 5;294(5540):30-1

Genomic and genetic definition of a functional human centromere. Schueler MG, Higgins AW, Rudd MK, Gustashaw K, Willard HF (USA)

The definition of centromeres of human chromosomes requires a complete genomic understanding of these regions. Toward this end, we report integration of physical mapping, genetic, and functional approaches, together with sequencing of selected regions, to define the centromere of the human X chromosome and to explore the evolution of sequences responsible for chromosome segregation. [PMID: 11588252]

-- Nat Genet. 1997 Apr;15(4):345-55; Comment in: Nat Genet. 1997 Apr;15(4):333-5.

Formation of de novo centromeres and construction of first-generation human artificial microchromosomes. Harrington JJ, Van Bokkelen G, Mays RW, Gustashaw K, Willard HF (USA)

We have combined long synthetic arrays of alpha satellite DNA with telomeric DNA and genomic DNA to generate artificial chromosomes in human HT1080 cells. This first-generation system for the construction of human artificial chromosomes should be suitable for dissecting the sequence requirements of human centromeres, as well as developing constructs useful for therapeutic applications. [PMID: 9090378]

-- J Cell Biol. 2002 Dec 9;159(5):765-75. Epub 2002 Dec 02

CENP-B box is required for de novo centromere chromatin assembly on human alphoid DNA. Ohzeki J, Nakano M, Okada T, Masumoto H. (Japan)

Centromere protein (CENP) B boxes, recognition sequences of CENP-B, appear at regular intervals in human centromeric alpha-satellite DNA (alphoid DNA). In this study, to determine whether information carried by the primary sequence of alphoid DNA is involved in assembly of functional human centromeres, we created four kinds of synthetic repetitive sequences. To the best of our knowledge, this is the first reported evidence of a functional molecular link between a centromere-specific DNA sequence and centromeric chromatin assembly in humans. [PMID: 12460987]

-- Genomics. 2002 Mar;79(3):297-304

Efficiency of de novo centromere formation in human artificial chromosomes. Mejia JE, Alazami A, Willmott A, Marschall P, Levy E, Earnshaw WC, Larin Z. (UK)

In a comparative study, we show that human artificial chromosome (HAC) vectors based on alpha-satellite (alphoid) DNA from chromosome 17 but not the Y chromosome regularly form HACs in HT1080 human cells. [PMID: 11863359]

-- Mol Ther. 2002 Jun;5(6):798-805

Alpha-satellite DNA and vector composition influence rates of human artificial chromosome formation. Grimes BR, Rhoades AA, Willard HF. (USA)

Human artificial chromosomes (HACs) have been proposed as a new class of potential gene transfer and gene therapy vector. HACs can be formed when bacterial cloning vectors containing alpha-satellite DNA are transfected into cultured human cells. The data presented here have a significant impact on the design of future HAC vectors that have potential to be developed for therapeutic applications and as tools for investigating human chromosome structure and function. (c)2002 Elsevier Science (USA). [PMID: 12027565]

-- Genomics. 1990 Aug;7(4):607-13

Pulsed-field gel analysis of alpha-satellite DNA at the human X chromosome centromere: high-frequency polymorphisms and array size estimate. Mahtani MM, Willard HF. (USA)

Using pulsed-field gel analysis (PFGE), we have characterized the large array of alpha-satellite DNA located in the centromeric region of the human X chromosome. PMID: [1974881]

-- Curr Opin Genet Dev. 1998 Apr;8(2):219-25

Centromeres: the missing link in the development of human artificial chromosomes. Willard HF. (USA)

Successful construction of artificial chromosomes is an important step for studies to elucidate the DNA elements necessary for chromosome structure and function. A roadblock to developing a tractable system in multicellular organisms, including humans, is the poorly understood nature of centromeres. The development of human artificial chromosome systems should facilitate investigation of the DNA and chromatin requirements for active centromere assembly. [PMID: 9610413]

-- Hum Mol Genet. 1998;7(10):1635-40

Engineering mammalian chromosomes. Grimes B, Cooke H. (UK)

Construction of a mammalian artificial chromosome (MAC) will develop our understanding of the requirements for normal chromosome maintenance, replication and segregation while offering the capacity for introducing genes into cells. These results demonstrate for the first time that alpha-satellite DNA can seed de novo centromeres in human cells, indicating that this repetitive sequence family plays an important role in centromere function. The stability of these MACs suggests that they have potential to be developed as gene delivery vectors. [PMID: 9735385]

-- Curr Opin Mol Ther. 2001 Apr;3(2):125-32

Satellite DNA-based artificial chromosomes for use in gene therapy. Hadlaczky G. (Hungary)

Satellite DNA-based artificial chromosomes (SATACs) can be made by induced de novo chromosome formation in cells of different mammalian species. These artificially generated accessory chromosomes are composed of predictable DNA sequences and they contain defined genetic information. Prototype human SATACs have been successfully constructed in different cell types ... [PMID: 11338924]

-- Genet Med. 2004 Mar-Apr;6(2):81-9

Detection of deletions in de novo "balanced" chromosome rearrangements: further evidence for their role in phenotypic abnormalities. Astbury C, Christ LA, Aughton DJ, Cassidy SB, Kumar A, Eichler EE, Schwartz S. (USA)

The purpose of this study was to test the hypothesis that deletions of varying sizes in de novo apparently balanced chromosome rearrangements are a significant cause of phenotypic abnormalities. A total of fifteen patients, with seemingly balanced de novo rearrangements by routine cytogenetic analysis but with phenotypic anomalies, were systematically analyzed. We characterized the breakpoints in these fifteen cases (two of which were ascertained prenatally), using a combination of high-resolution GTG-banding, fluorescence in situ hybridization (FISH) with bacterial artificial chromosomes (BACs), and data from the Human Genome Project.

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