DNA walking PCR Service

For the few species whose genomes have been sequenced (e.g., human, mouse, nematode, rice, Arabidopsis, etc.), the sequences flanking the known sequences of a particular species can be easily found in databases. However, for most organisms, the way to know the DNA sequences flanking a known region without knowing their genome sequence is to use DNA walking techniques. DNA walking is an important molecular biology research technique that can be used to obtain unknown sequences adjacent to known sequences efficiently.

There are more than a dozen PCR-based DNA walking techniques, and Lifeasible can provide you with TAIL-PCR (thermal asymmetric interlaced PCR). It uses specific nested primers, combined with concatenated primers to fish for flanking sequences.

Basic principle

We design three nested specific primers (SP1, SP2, SP3) using known sequences next to the target sequence and use each of them in combination with a short, random, degenerate primer (AD) with a low Tm value. An asymmetric temperature cycle is then designed according to the differences in primer length and specificity, and the specific product is amplified by a graded reaction. TAIL-PCR technology has the advantages of simplicity, speed, high specificity, accurate and reliable reaction products, and good reproducibility.

Service flow

Delivery contents

• Experiment report: primer information, experiment process, PCR product gel picture, target sequence.
• Sequencing results: all sequencing files related to the experiment.

Applications

• Acquisition of critical regulatory genes in humans, animals, and plants in successive steps based on known genes or molecular markers can be used to study the regulation of expression of structural genes. For example, the isolation of cloned promoters and the study of their functions.
• Stepwise access to non-conserved regions of genes in new species, thus obtaining complete gene sequences.
• Identification of insertion sites of T-DNA or transposons, identification of insertion sites of exogenous genes resulting from transgenic techniques such as the gene gun transgenic method.
• For gap filling in chromosome sequencing to obtain the complete genome sequence.
• Used for segment lapping of artificial chromosomes PAC, YAC, and BAC.

Advantages

• We have rich experimental experience in designing simple and degenerate primers.
• It can be connected with our transgenic service, detection service, protein service, etc., and the whole experiment can be completed smoothly and quickly.

Lifeasible provides DNA walking technology services to help you identify the order in which specific DNA fragments have been cloned and flanked. Please feel free to contact us to submit your requirements.