A rapid DNA cloning system is a research interest of many scientists. TA cloning is one of the methods used for the cloning of PCR-amplified DNA molecules. The TA cloning method is a convenient and labor-saving replacement to traditional, restriction enzyme-mediated cloning strategies. A T-vector called pBlueskript ΙΙ SK-1 with the lethal gene ccdB was designed to construct a positive selection...

Multiple gene silencing is being required to target and tangle metabolic pathways in eukaryotes and researchers have to develop a subtle method for construction of RNA interference (RNAi) cassettes. Although, several vectors have been developed due to different screening and cloning strategies but still some potential limitations remain to be dissolved. Here, we worked out a simple cloning stra...

multiple gene silencing is being required to target and tangle metabolic pathways in eukaryotes and researchers have to develop a subtle method for construction of rna interference (rnai) cassettes. although, several vectors have been developed due to different screening and cloning strategies but still some potential limitations remain to be dissolved. here, we worked out a simple cloning stra...

Direct cloning of PCR fragments by TA cloning or blunt end ligation are two simple methods which would greatly benefit high-throughput (HTP) cloning constructions if the efficiency can be improved. In this study, we have developed a ribosomal binding site (RBS) switching strategy for direct cloning of PCR fragments. RBS is an A/G rich region upstream of the translational start codon and is esse...

Molecular cloning methods based on primer and overlap-extension PCR are widely used due to their simplicity, reliability, low cost and high efficiency. In this article, an efficient mega primer-mediated (MP) cloning strategy for chimaeragenesis and long DNA fragment insertion is presented. MP cloning is a seamless, restriction/ligation-independent method that requires only three steps: (i) the ...

We describe a cloning strategy, third irrelevant enzyme site (TIES), based on three-component assembly ligation reactions to overcome seven types of problematical cloning reactions. Implementation of the TIES strategy has made it possible to obtain the desired recombinant where the corresponding approach using conventional recombinant DNA methodologies and two-component ligations failed to give...

We have developed an efficient strategy for cloning of PCR products that contain an unknown region flanked by a known sequence. As with ligation-independent cloning, the strategy is based on homology between sequences present in both the vector and the insert. However, in contrast to ligation-independent cloning, the cloning vector has homology with only one of the two primers used for amplific...

Long fragment cloning is a challenge for its difficulty in accurate amplifying and tendency to get unwanted mutation. Here we discuss Restriction-based Multiple-fragment Assembly Strategy's advantages and limitations. In this strategy, rather than PCR amplifying the entire coding sequence (CDS) at one time, we amplified and sequenced smaller fragments which are shorter than 1.5kb spanning the C...

A scheme of quantum feedback control with an optimal cloning machine is proposed. The design of quantum feedback control algorithms is separated into a state recognition strategy, which gives “on-off” signal to the actuator through recognizing some copies from cloning machine, and a feedback (control) strategy through feeding back the another copies of cloning machine. Precise feedback is aband...