Congenital Abnormalities ; genetics ; Gene Dosage ; Genome, Human ; Humans

Multiple system atrophy (MSA) is a progressive neurodegenerative disorder. Widespread presence of glial cytoplasmic inclusions is the neuropathologic hallmark of MSA. The disease has long been considered as a sporadic disorder. However, in recent years, a few familial cases of MSA have been reported, and researches have verified certain genetic variants could increase the risk of MSA. These indicated genetic factors may play an imported role in the pathogenesis of MSA. In this review, the emerging evidence in favor of genetic players in MSA is discussed.
Animals ; Gene Dosage ; Genetic Research ; Humans ; Multiple System Atrophy ; genetics

OBJECTIVETo examine the feasibility of linking operons in tandem to enhance expression of heterologous genes in Escherichia coli (E. coli) and clarify the potential control mechanism of the total plasmid DNA amount in each host cell.

METHODSTwo series of expression plasmids, CW11 and CW12, containing 1 to 4 and 1 to 3 heterologous gene operon(s) respectively, were constructed. The molecular size of the CW11 series varied from 5.47 kb to 12.26 kb in 2.25 kb increments. The CW12 series varied from 5.40 kb to 9.72 kb in 2.16 kb increments. The expression level of desired protein was assayed by SDS-PAGE and laser density scanning. Plasmid copy number was determined by incorporation with (3)H-thymidine ((3)H-TdR).

RESULTSNo influence of the tandem-joined operons on host growth and plasmid stability was observed. Upon induction, the desired protein accumulations in the CW11 series were 44.9% +/- 3.9%, 51.3% +/- 4.1%, 54.8% +/- 3.3% and 58.2% +/- 3.4% of total cell protein. In the CW12 series, the yields were 32.2% +/- 5.0%, 42.8% +/- 4.1% and 46.9% +/- 4.0% of total cell protein. As size increased, the plasmid copy number decreased, but target gene dosage increased significantly (P < 0.01). Further calculation showed that the total amount of plasmid DNA per cell was not significantly different in each series (P > 0.05) and restricted to some extent.

CONCLUSIONSIncreasing the target gene dosage by tandem linking of operons may enhance the expression level of a desired protein. Although the size (kb) and the copy number of each plasmid are negatively interrelated, for certain plasmids in each series, their total DNA amount per cell seems to be a restricted constant for specific E. coli strains under identical incubation condition.

DNA ; analysis ; Escherichia coli ; genetics ; Gene Dosage ; Operon ; Plasmids

OBJECTIVETo investigate the correlation of the deleted azoospermia (DAZ) gene copy related to gr/gr and b2/b3 deletions in the AZFc region with male spermatogenic impairment.

METHODSThis study included 121 infertile men with different de- grees of spermatogenic impairment and 95 healthy donors from the sperm bank. Using PCR, PCR-RFLP, and Y chromosome specific sequence tagged sites (STS) , we analyzed the association of DAZ gene copy deletions related to gr/gr and b2/b3 deletions in the AZFc region with spermatogenic impairment.

RESULTSThere were 15 cases of gr/gr deletion (12. 40% ) and 6 cases of b2/b3 deletion (4.96%) in the infertility group as compared with 13 cases of gr/gr deletion (13.68%) and 1 case of b2/b3 deletion (1.05%) in the control. Analysis of the DAZ-specific single nucleotide variant (SNV) loci revealed 11 gr/gr-DAZI/DAZ2 deletions (9.09%), 4 gr/gr-DAZ3/DAZ4 deletions (3.31%), and 6 b2/b3-DAZ1/DAZ2 deletions (4.96%) in the infertile men in comparison with 3 gr/ gr-DAZ1/DAZ2 deletions (3.16%), 10 gr/gr-DAZ3/DAZ4 deletions (10.53%), and 1 b2/b3- DAZ3/DAZ4 deletion (1.05%) in the control.

CONCLUSIONPartial deletions of gr/gr and b2/b3 exist in both healthy men and male patients with different degrees of spermatogenic impairment and cannot be considered as a risk factor for spermatogenesis impairment. However, deletions of different DAZ duplicons in gr/gr and b2/b3 deletions have different effects on spermatogenesis. DAZ1/DAZ2 instead of DAZ3/DAZ4 deletions might be associated with spermatogenesis impairment.

Deleted in Azoospermia 1 Protein ; Gene Deletion ; Gene Dosage ; Humans ; Male ; RNA-Binding Proteins ; genetics ; Spermatogenesis ; genetics

OBJECTIVETo investigate the clinicopathological significance of chromosome 17 polysomy in breast cancer.

METHODSRetrospective study of 200 cases of breast cancer including 106 cases of invasive ductal carcinoma and 94 cases of in-situ carcinoma was performed by fluorescence in-situ hybridization (FISH) to explore the relationship between chromosome 17 polysomy and age, nuclear atypia, lymphatic metastasis, HER2 gene amplification and HER2 protein expression.

RESULTSTwenty-six percent (52/200) of chromosome 17 polysomy was detected in 200 cases of breast ductal carcinoma, all of which were invasive ductal carcinoma. Overall 52. 8% (52/180) of invasive ductal carcinoma cases showed chromosome 17 polysomy, which was correlated to HER2 gene amplification (P = 0.000) and HER-2 protein expression (P=0.000), and to HER2 expression combined with HER2 gene amplification (P=0.001). Chromosome 17 polysomy with or without HER2 gene amplification was also associated with high-grade nuclear atypia (P = 0.012 or P = 0.010) and lymphatic metastasis (P = 0.002 or P = 0.009 ). However, chromosome 17 polysomy with or without HER2 gene amplification was not correlative with the age of patients (P = 1. 000 or P = 0. 415).

CONCLUSIONChromosome 17 polysomy may be related to the nuclear atypia, metastasis, HER2 gene amplification of invasive ductal carcinoma and thus a worse prognosis of the patients.

Breast Neoplasms ; genetics ; pathology ; Carcinoma, Ductal ; genetics ; Chromosome Aberrations ; Chromosomes, Human, Pair 17 ; genetics ; Gene Amplification ; Gene Dosage ; Gene Expression Regulation, Neoplastic ; genetics ; Genes, erbB-2 ; genetics ; Humans

How to reduce immune response is an unprecedented challenge for rAAV gene medicine. Recent studies suggested that lowering dosage of the vector used could reduce immune response caused by rAAV gene medicine. Nevertheless, it would also decrease the transgene expression, leading to failure of gene treatment. It is therefore important to take appropriate steps to maintain high gene expression level and pharmacodynamic, while the dosage of rAAV used is reduced. Here, steps to enhancing gene therapy, such as optimization of the administration, reconstruction of the viral vector and selection of the promoter, are discussed in order to achieve maximum outcome.
Animals ; Dependovirus ; genetics ; immunology ; Dose-Response Relationship, Drug ; Gene Dosage ; Gene Transfer Techniques ; Genetic Therapy ; Genetic Vectors ; administration & dosage ; genetics ; immunology ; Humans ; Recombination, Genetic ; Transgenes ; genetics

Gene dosage determination is increasingly important for the study of both genome variation and rearrangement associated with complex diseases. Large genomic duplications and deletions are increasingly found as the causes. Methods such as PCR or sequencing are usually qualitative rather than quantitative. Thus, these methods can not detect large genomic duplications or deletions. Therefore, searching for a gene dosage method which is reliable, sensitive and high-throughput becomes imperative. Many high-performance technologies have been developed for gene dosage analyses in the recent years. There are generally three categories of methods including cytogenetic, Southern or dot blotting, or PCR amplification. Recent development in these techniques have been introduced and discussed in this review, which will help people to choose a suitable method for different research.
Blotting, Southern ; methods ; Cytogenetic Analysis ; methods ; Gene Deletion ; Gene Dosage ; genetics ; Gene Duplication ; Humans ; Polymerase Chain Reaction ; methods

The completion of many malaria parasite genomes provides great opportunities for genomewide characterization of gene expression and high-throughput genotyping. Substantial progress in malaria genomics and genotyping has been made recently, particularly the development of various microarray platforms for large-scale characterization of the Plasmodium falciparum genome. Microarray has been used for gene expression analysis, detection of single nucleotide polymorphism (SNP) and copy number variation (CNV), characterization of chromatin modifications, and other applications. Here we discuss some recent advances in genetic mapping and genomic studies of malaria parasites, focusing on the use of high-throughput arrays for the detection of SNP and CNV in the P. falciparum genome. Strategies for genetic mapping of malaria traits are also discussed.
Animals ; *Chromosome Mapping ; DNA, Protozoan/*genetics ; Gene Dosage ; *Microarray Analysis ; Plasmodium falciparum/*genetics ; Polymorphism, Genetic

In order to improve the fermentation potency of phytase in recombinant host and decrease the production cost, the pichia expression vector pGAPZalpha-A was modified by introduction of an AOX1 promoter from vector pPIC9 and the resulted vector pAOXZalpha is an methanol induced vector. After that, a phytase gene appA-m was cloned into pAOXZalpha to construct the recombinant vector pAOXZalpha-appA-m. The recombinant Pichia pastoris 74#, which already contains one copy of appA-m and its fermentation potency exceeded 7.5 x 10(6) IU/mL, was used as the host strain for the transformation of pAOXZalpha-appA-m. The Pichia pastoris transformants were gained by electroporation. PCR results indicated that the appA-m expression box has integrated into the genome of Pichia pastoris and the original construction of phytase gene has not changed. SDS-PAGE analysis revealed that phytase was overexpressed and secreted into the medium supernatant. Recombinants with high expression level were screened and used for fermentation. In 5L fermentor, the expression level of phytase protein achieved 4 mg/mL and the phytase activity (fermentation potency) exceeded 1.2 x 10(7) IU/mL, which was about 1.6-fold compared with that of the host strain 74#. Moreover, the improved recombinant Pichia pastoris is excellent at expression stability and heredity stability.
6-Phytase ; genetics ; Fermentation ; Gene Dosage ; Pichia ; genetics ; Plasmids ; Polymerase Chain Reaction ; Recombination, Genetic

OBJECTIVETo establish a primed in situ labeling (PRINS) technique which can be more effective in detection of single copy gene.

METHODSOn the basis of traditional PRINS, new reagents and procedures, such as TaqStart antibody, four primers of the sex determining region Y (SRY) gene and TSA(TM) Biotin System were included in detection of the SRY gene. Meanwhile, fluorescence in situ hybridization(FISH) to detect the SRY gene was used as control.

RESULTSFifty metaphases were scored. PRINS labeling showed signals for the SRY on the Y chromosome at band Yp11.3 in all metaphases. These signals were as distinct as that from results of FISH.

CONCLUSIONThis improved method is ideal for rapidly localizing single copy genes and small DNA segments. And PRINS is a cost- and time-effective alternative to FISH.

Gene Dosage ; Genes, sry ; genetics ; Humans ; In Situ Hybridization, Fluorescence ; Male ; Metaphase ; genetics ; Primed In Situ Labeling ; methods