Animal cloning has been achieved in many species by transplanting differentiated cell nuclei to unfertilized oocytes. However, the low efficiencies of cloning have remained an unresolved issue. Here we find that the combination of two small molecules, trichostatin A (TSA) and vitamin C (VC), under culture condition with bovine serum albumin deionized by ion-exchange resins, dramatically improve...

Somatic cell nuclear transfer (SCNT) has been used for the cloning of various mammals. However, the rates of successful, healthy birth are generally poor. To improve cloning efficiency, we report the utilization of an 'autologous SCNT' cloning technique in which the somatic nucleus of a female bovine donor is transferred to its own enucleated oocyte recovered by ovum pick up, in contrast to the...

As the demand for cloned embryos and offspring increases, the need arises for the development of nuclear transfer procedures that are improved in both efficiency and ease of operation. Here, we describe a novel zona-free cloning method that doubles the throughput in cloned bovine embryo production over current procedures and generates viable offspring with the same efficiency. Elements of the p...

BACKGROUND One of the major challenges for membrane protein structural genomics is establishing high-throughput cloning and expression screening methods to obtain enough purified protein in a homogeneous preparation for structural and functional studies. Here a series of ligation independent cloning based vectors were constructed to address this challenge. RESULTS The feasibility of these vec...

Imprinting disorders induced by somatic cell nuclear transfer (SCNT) usually lead to the abnormalities of cloned animals and low cloning efficiency. Histone deacetylase inhibitors have been shown to improve gene expression, genomic methylation reprogramming and the development of cloned embryos, however, the imprinting statuses in these treated embryos and during their subsequent development re...

Current standard cloning methods based on the use of restriction enzymes and ligase are very versatile, but are not well suited for high-throughput cloning projects or for assembly of many DNA fragments from several parental plasmids in a single step. We have previously reported the development of an efficient cloning method based on the use of type IIs restriction enzymes and restriction-ligat...

A couple of DNA ligation-independent cloning (LIC) methods have been reported to meet various requirements in metabolic engineering and synthetic biology. The principle of LIC is the assembly of multiple overlapping DNA fragments by single-stranded (ss) DNA overlaps annealing. Here we present a method to generate single-stranded DNA overlaps based on Nicking Endonucleases (NEases) for LIC, the ...

The precise assembly of defined DNA sequences into plasmids is an essential task in bioscience research. While a number of molecular cloning techniques have been developed, many methods require specialized expensive reagents or laborious experimental procedure. Not surprisingly, conventional cloning techniques based on restriction digestion and ligation are still commonly used in routine DNA cl...

Dolly the sheep cloning “the first cloning process” was derived by somatic cell nuclear transfer and it had reported that the clone got premature senescence as a result of continues telomere shortening and ongoing decrease in telomerase activity by time; thus majority of recent researches is directed to use other types of donor cells to be transferred into the recipient enucleated oocyte to mai...

BACKGROUND Cloning vectors capable of retroviral transduction have enabled stable gene overexpression in numerous mitotic cell lines. However, the relatively small number of feasible restriction enzyme sequences in their cloning sites can hinder successful generation of overexpression constructs if these sequences are also present in the target cDNA insert. RESULTS Utilizing ligation-independ...