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Nature Methods Jul 2013Tetrad analysis has been a gold-standard genetic technique for several decades. Unfortunately, the need to manually isolate, disrupt and space tetrads has relegated its...
Tetrad analysis has been a gold-standard genetic technique for several decades. Unfortunately, the need to manually isolate, disrupt and space tetrads has relegated its application to small-scale studies and limited its integration with high-throughput DNA sequencing technologies. We have developed a rapid, high-throughput method, called barcode-enabled sequencing of tetrads (BEST), that uses (i) a meiosis-specific GFP fusion protein to isolate tetrads by FACS and (ii) molecular barcodes that are read during genotyping to identify spores derived from the same tetrad. Maintaining tetrad information allows accurate inference of missing genetic markers and full genotypes of missing (and presumably nonviable) individuals. An individual researcher was able to isolate over 3,000 yeast tetrads in 3 h, an output equivalent to that of almost 1 month of manual dissection. BEST is transferable to other microorganisms for which meiotic mapping is significantly more laborious.
Topics: Algorithms; Chromosome Mapping; DNA, Fungal; Genetic Markers; High-Throughput Nucleotide Sequencing; Meiosis; Saccharomyces cerevisiae
PubMed: 23666411
DOI: 10.1038/nmeth.2479 -
Zhejiang Da Xue Xue Bao. Yi Xue Ban =... Aug 2021Neonatal genetic disease is currently screened mainly based on metabolite biochemical technology. The false positive rate of biochemical screening technology is...
Neonatal genetic disease is currently screened mainly based on metabolite biochemical technology. The false positive rate of biochemical screening technology is relatively high, and there are certain false negatives, and only few types of diseases can be screened. The genetic techniques have been gradually used for neonatal genetic disease screening in recent years. Gene detection technology includes quantitative PCR (qPCR) and high-throughput sequencing. High-throughput sequencing includes gene panel sequencing, whole-exome sequencing and whole-genome sequencing. At present, qPCR and gene panel sequencing are the main technologies to be used for newborn genetic disease screening. Genetic screening diseases range from single disease such as hearing loss, spinal muscular atrophy and severe combined immunodeficiency to multiple diseases. Besides standards and guidelines for the interpretation of sequence variants proposed by the American College of Medical Genetics and Genomics and the Association for Molecular Pathology in 2015, the interpretation of genetic screening results should also consider biochemical results and other results. The development of newborn genetic screening needs to follow ethical principles, including the ethics of newborn genetic screening as a public health project, the privacy ethics of newborns and their family members, and the ethics of bioinformatics. The development of newborn genetic screening will enable more patients with inherited diseases to receive early diagnosis and treatment and improve their prognosis, which is a milestone in the field of neonatal screening.
Topics: Genetic Techniques; Genetic Testing; Genomics; Humans; Infant, Newborn; Neonatal Screening; United States; Exome Sequencing
PubMed: 34704410
DOI: 10.3724/zdxbyxb-2021-0288 -
Cold Spring Harbor Perspectives in... Mar 2019Long double-stranded RNAs (dsRNAs) are abundantly expressed in animals, in which they frequently occur in introns and 3' untranslated regions of mRNAs. Functions of... (Review)
Review
Long double-stranded RNAs (dsRNAs) are abundantly expressed in animals, in which they frequently occur in introns and 3' untranslated regions of mRNAs. Functions of long, cellular dsRNAs are poorly understood, although deficiencies in adenosine deaminases that act on RNA, or ADARs, promote their recognition as viral dsRNA and an aberrant immune response. Diverse dsRNA-binding proteins bind cellular dsRNAs, hinting at additional roles. Understanding these roles is facilitated by mapping the genomic locations that express dsRNA in various tissues and organisms. ADAR editing provides a signature of dsRNA structure in cellular transcripts. In this review, we detail approaches to map ADAR editing sites and dsRNAs genome-wide, with particular focus on high-throughput sequencing methods and considerations for their successful application to the detection of editing sites and dsRNAs.
Topics: Adenosine Deaminase; Animals; Genetic Techniques; Humans; RNA, Double-Stranded
PubMed: 30824577
DOI: 10.1101/cshperspect.a035352 -
Methods in Molecular Biology (Clifton,... 2011Ascidians, such as Ciona, are invertebrate chordates with simple embryonic body plans and small, relatively non-redundant genomes. Ciona genetics is in its infancy...
Ascidians, such as Ciona, are invertebrate chordates with simple embryonic body plans and small, relatively non-redundant genomes. Ciona genetics is in its infancy compared to many other model systems, but it provides a powerful method for studying this important vertebrate outgroup. Here we give basic methods for genetic analysis of Ciona, including protocols for controlled crosses both by natural spawning and by the surgical isolation of gametes; the identification and propagation of mutant lines; and strategies for positional cloning.
Topics: Animals; Chromosome Mapping; Cloning, Molecular; Cryopreservation; DNA; DNA Mutational Analysis; Female; Fertilization in Vitro; Genetic Techniques; Hybridization, Genetic; Larva; Male; Microinjections; Mutation; Polymerase Chain Reaction; Polymorphism, Single Nucleotide; Urochordata
PubMed: 21805273
DOI: 10.1007/978-1-61779-210-6_15 -
Molecular Biology Reports Jul 2022In recent years, the number of tools and techniques that enable genetic material to be added, removed or altered at specific locations in the genome has increased...
In recent years, the number of tools and techniques that enable genetic material to be added, removed or altered at specific locations in the genome has increased significantly. The objective is to know the structure of genomes, the function of genes and improve gene therapy.In this work we intend to explain the functioning of the CRISPR/Cas9 (Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR associated protein 9) and the advantages that this technique may have compared to previously developed techniques, such as RNA interference (RNAi), Zinc Finger Nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) in gene and genome editing.We will start with the story of the discovery, then its biological function in the adaptive immune system of bacteria against bacteriophage attack, and ending with a description of the mechanism of action and its use in gene editing. We will also discuss other Cas enzymes with great potential for use in genome editing as an alternative to Cas9.CRISPR/Cas9 is a simple, inexpensive, and effective technique for gene editing with multiple applications from the development of functional genomics and epigenetics. This technique will, in the near future, have great applications in the development of cell models for use in medical and pharmaceutical processes, in targeted therapy, and improvement of agricultural and environmental species.
Topics: CRISPR-Cas Systems; Epigenesis, Genetic; Gene Editing; Genetic Therapy; Genome
PubMed: 35716290
DOI: 10.1007/s11033-022-07442-w -
BioMed Research International 2015Genetics is the study of heredity, which means the study of genes and factors related to all aspects of genes. The scientific history of genetics began with the works of... (Review)
Review
Genetics is the study of heredity, which means the study of genes and factors related to all aspects of genes. The scientific history of genetics began with the works of Gregor Mendel in the mid-19th century. Prior to Mendel, genetics was primarily theoretical whilst, after Mendel, the science of genetics was broadened to include experimental genetics. Developments in all fields of genetics and genetic technology in the first half of the 20th century provided a basis for the later developments. In the second half of the 20th century, the molecular background of genetics has become more understandable. Rapid technological advancements, followed by the completion of Human Genome Project, have contributed a great deal to the knowledge of genetic factors and their impact on human life and diseases. Currently, more than 1800 disease genes have been identified, more than 2000 genetic tests have become available, and in conjunction with this at least 350 biotechnology-based products have been released onto the market. Novel technologies, particularly next generation sequencing, have dramatically accelerated the pace of biological research, while at the same time increasing expectations. In this paper, a brief summary of genetic history with short explanations of most popular genetic techniques is given.
Topics: Genetic Techniques; Genetics; History, 20th Century; History, 21st Century; Humans
PubMed: 25874212
DOI: 10.1155/2015/461524 -
Journal of Medical Ethics Jun 2005The diagnosis of bipolar disorder has been linked to giftedness of various sorts and this raises a special problem in that it is likely that the condition has a genetic... (Review)
Review
The diagnosis of bipolar disorder has been linked to giftedness of various sorts and this raises a special problem in that it is likely that the condition has a genetic basis. Therefore it seems possible that in the near future we will be able to detect and eliminate the gene predisposing to the disorder. This may mean, however, that, as a society, we lose the associated gifts. We might then face a difficult decision either way in that it is unclear that we are preventing an unalloyed bad when we diagnose and eliminate bipolar disorder through prenatal genetic testing and yet if we allow the individual to be born we are condemning that person to being an unwitting sacrifice in that they might well suffer considerable net distress as a result of our need to keep our gene pool enriched in the relevant way.
Topics: Abortion, Induced; Bipolar Disorder; Creativity; Culture; Ethical Theory; Genetic Techniques; Genome, Human; Harm Reduction; Humans; Prenatal Diagnosis; Value of Life
PubMed: 15923479
DOI: 10.1136/jme.2004.009613 -
Microbial Cell Factories Oct 2017Filamentous fungi have been of great interest because of their excellent ability as cell factories to manufacture useful products for human beings. The development of... (Review)
Review
Filamentous fungi have been of great interest because of their excellent ability as cell factories to manufacture useful products for human beings. The development of genetic transformation techniques is a precondition that enables scientists to target and modify genes efficiently and may reveal the function of target genes. The method to deliver foreign nucleic acid into cells is the sticking point for fungal genome modification. Up to date, there are some general methods of genetic transformation for fungi, including protoplast-mediated transformation, Agrobacterium-mediated transformation, electroporation, biolistic method and shock-wave-mediated transformation. This article reviews basic protocols and principles of these transformation methods, as well as their advantages and disadvantages.
Topics: Agrobacterium; Biolistics; Electroporation; Fungi; Gene Transfer Techniques; Genetic Techniques; Genome, Fungal; Protoplasts; Transformation, Genetic
PubMed: 28974205
DOI: 10.1186/s12934-017-0785-7 -
Cold Spring Harbor Perspectives in... Oct 2014The skin is composed of a variety of cell types expressing specific molecules and possessing different properties that facilitate the complex interactions and... (Review)
Review
The skin is composed of a variety of cell types expressing specific molecules and possessing different properties that facilitate the complex interactions and intercellular communication essential for maintaining the structural integrity of the skin. Importantly, a single mutation in one of these molecules can disrupt the entire organization and function of these essential networks, leading to cell separation, blistering, and other striking phenotypes observed in inherited skin diseases. Over the past several decades, the genetic basis of many monogenic skin diseases has been elucidated using classical genetic techniques. Importantly, the findings from these studies has shed light onto the many classes of molecules and essential genetic as well as molecular interactions that lend the skin its rigid, yet flexible properties. With the advent of the human genome project, next-generation sequencing techniques, as well as several other recently developed methods, tremendous progress has been made in dissecting the genetic architecture of complex, non-Mendelian skin diseases.
Topics: Chromosomal Position Effects; Cloning, Molecular; DNA Methylation; Diseases in Twins; Gene Expression Profiling; Genetic Techniques; Genome-Wide Association Study; Humans; MicroRNAs; Mosaicism; Mutation; Oligonucleotide Array Sequence Analysis; Phenotype; Skin Diseases, Genetic; Uniparental Disomy
PubMed: 25274756
DOI: 10.1101/cshperspect.a015172 -
Journal of Cellular and Molecular... 2006Brain structure and function are determined in part through experience and in part through our inherited genes. A powerful approach for unravelling the balance between... (Review)
Review
Brain structure and function are determined in part through experience and in part through our inherited genes. A powerful approach for unravelling the balance between activity-dependent neuronal plasticity and genetic programs is to directly manipulate the genome. Such molecular genetic studies have been greatly aided by the remarkable progress of large-scale genome sequencing efforts. Sophisticated mouse genetic manipulations allow targeted point-mutations, deletions and additions to the mouse genome. These can be regulated through inducible promoters expressing in genetically specified neuronal cell types. However, despite significant progress it remains difficult to target specific brain regions through transgenesis alone. Recent work suggests that transduction vectors, like lentiviruses and adeno-associated viruses, may provide suitable additional tools for localized and controlled genetic manipulation. Furthermore, studies with such vectors may aid the development of human genetic therapies for brain diseases.
Topics: Animals; Brain; Dependovirus; Gene Expression Regulation; Gene Targeting; Gene Transfer Techniques; Genetic Techniques; Genetic Vectors; Genome; Lentivirus; Mice; Mice, Transgenic
PubMed: 16796803
DOI: 10.1111/j.1582-4934.2006.tb00403.x