Site-Directed Mutagenesis

Site-Directed mutation is one of the most important techniques used in protein engineering, which can make specific and intentional changes to the DNA sequence of a gene and any gene products. It can be used to elucidate the mechanism of gene regulation and study the relationship between protein structure and functional site-directed mutation. In order to maintain the integrity of original sequences, the efficiency of your experiment can be increased by providing a single base or multiple bases in your mutant template plasmid, including deletions, insertions, and site mutations. Our company provides you high quality mutant products in the shortest time.

1. Short TAT: common sequence can be delivered to you within 5 working days;

2. Vast operating region: if the distance between two point mutations is <30bp, we deal it as one;

3. High quality: we has rich experiences in solving kinds of repetitive sequences, including hairpin structure, high GC, poly structure and other complex sequences perfectly;

4. Mutations on Large DNA Constructs: Our technology is optimized to introduce point mutations, insertion mutations, and deletion mutations on constructs as large as 12 kb (including target gene and its vector);

5. One-stop solutions: We offer comprehensive upstream and downstream services, including template DNA sequencing, de novo gene synthesis, Express Cloning into free expression vectors, customized vector construction, and protein expression and purification;

6. Most Competitive Prices starting at $66/mutant.

1. What materials and information do I need to provide the company for site-directed mutations?

Answer

Materials needed to be provided:

A template plasmid 4μl or a bacterial solution which contains a template plasmid with a plasmid or germ liquid of the target carrier.

Information you need to provide:

a) template gene name and sequence (pre-mutation sequence), template sequence of sequencing results;

b) the vector information of template sequence: full name, size and resistance of the carrier;

c) gene name and sequence after mutation;

d) target carrier information: full name, size and resistance of the carrier. If there is non-commercial carrier, need to provide the whole sequence of vectors; 

2. If I want to replace the target vector plasmid, how to charge?

Answer

In addition to the site-directed mutation, you also need pay the fee of sub-cloning.

3. May I introduce point mutation in the genome with genomic editing techniques?

Answer

Yes, you need donor DNA with the target homologous recombination arm and the desired point mutation, which is a plasmid or a single-stranded oligonucleotide.

4. Is it possible to get large numbers of colonies from PCR product after performing site directed mutagenesis on ampicillin containing plates?

Answer

Large colony counts after site directed mutagenesis are somewhat unusual. In general, a variation in colony numbers between 0-50 after transforming DH5a cells can be got.

5. What is the maximum deletion size applicable to achieve in site-directed mutagenesis?

Answer

In theory there is no limit to the deletion size of site-directed mutagenesis. Of course it is important that the deletions do not involve regions of the plasmid essential for vector replication and maintenance (e,g replication origin, antibiotic resistance).

6. Is it necessary to use specific plasmids as a template for gene mutations?

Answer

No specific plasmid is needed. Just make sure extension time is appropriate for the size of the plasmid.

7. What is the effect of purification method of primers on site directed mutagenesis?

Answer

Yes, the purpose of primer purification is to avoid the side effect of the presence of unwanted primer in PCR reactions. So for mutagenesis experiments, it's better to use the purified primers in order to improve the yield of the mutagenesis.

8. What is the maximum number of nucleotides that can be inserted between two nucleotides through a site directed mutations?

Answer

In theory there is no limit as long as you have the way to generate the suitable DNA fragment bearing homologies with the plasmid template at its ends for efficient annealing.

9. What is the most appropriate substitute amino acid for histidine in site directed mutagenisis?

Answer

It depends on your aims. Most common for abolishing catalytic function is the change to alanine. Some people want to mimic phosphorylation of serine so they put aspartate, while some want to increase or decrease the binding pocket size so they can substitute for bulky residues (W, I) or smaller residues (G, A).