Synthesis of DNA fragments in yeast by one-step assembly of overlapping oligonucleotides

نویسنده

  • Daniel G. Gibson
چکیده

Here it is demonstrated that the yeast Saccharomyces cerevisiae can take up and assemble at least 38 overlapping single-stranded oligonucleotides and a linear double-stranded vector in one transformation event. These oligonucleotides can overlap by as few as 20 bp, and can be as long as 200 nucleotides in length. This straightforward scheme for assembling chemically-synthesized oligonucleotides could be a useful tool for building synthetic DNA molecules.

برای دانلود رایگان متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید

ثبت نام

اگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید

منابع مشابه

One-step assembly in yeast of 25 overlapping DNA fragments to form a complete synthetic Mycoplasma genitalium genome.

We previously reported assembly and cloning of the synthetic Mycoplasma genitalium JCVI-1.0 genome in the yeast Saccharomyces cerevisiae by recombination of six overlapping DNA fragments to produce a 592-kb circle. Here we extend this approach by demonstrating assembly of the synthetic genome from 25 overlapping fragments in a single step. The use of yeast recombination greatly simplifies the a...

متن کامل

Specific Inhibition of the Expression of the Promyelocytic Leukemia (PML) Protein by Anti-Sense Oligonucleotides

In the present study, using anti-sense oligonucleotides the inhibition of expression of the PML protein hasbeen investigated. The anti-sense oligonucleotides were designed against the translation initiation site ofthe PML gene, and their effects were investigated on cellular growth and DNA synthesis. Incubation of normalhuman fibroblast cells with the anti-sense oligonucleotid...

متن کامل

Synthesis and Expression of Modified bFGF Gene in Escherichia coli Cells

A new strategy for construction of synthetic gene encoding human basic fibroblast growth factor comprising DNA annealing-ligation and augmentation by polymerase chain reaction was introduced. The sequence of the gene and corresponding amino acid chain were modified in order to increase stability of the protein. First, 300 bp and 160 bp fragments of the gene were assembled from 18 oligonucleotid...

متن کامل

In Situ Overlap and Sequence Synthesis During DNA Assembly

Modern cloning methods are independent from restriction enzyme recognition sites. However, nearly all current cloning methods still require the introduction of overlaps by PCR, which can introduce undesired mutations. Here, we investigated whether overlaps needed for DNA assembly can be synthesized in situ and we tested if the de novo synthesis of sequences can be simultaneously combined with t...

متن کامل

Varicella Zoster Virus (VZV) Origin-Dependent Plasmid Replication in the Presence of the Four Overlapping Cosmids Comprising the Complete Genome of VZV

The Varicella-Zoster Virus (VZV) genome contains both cis-acting and trans-acting elements, which are important in viral DNA replication. The cis-acting elements consist of two copies of oriS, and the trans-acting elements are those genes whose products are required for virus DNA replication. It has been shown that each of the seven genes required for ori-dependent DNA synthesis of Herpes Simpl...

متن کامل

ذخیره در منابع من


  با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید

عنوان ژورنال:

دوره 37  شماره 

صفحات  -

تاریخ انتشار 2009