DNA (Deoxyribonucleic Acid) library and method for constructing transcription activator like effector nuclease plasmids

Abstract

The invention discloses a DNA (Deoxyribonucleic Acid) library and a method for constructing transcription activator like effector nuclease plasmids. A Tale connecting cell library comprises 16 groups*n double-base identifying modules and 4 groups*n single-base identifying module; and a first viscous tail end and a second viscous tail end which are formed by enzyme digestion of two restriction enzymes are arranged on two tail ends of each double-base identifying module or each single-base identifying module; the DNA library comprises plasmids which are as many as connecting units of the Tale connecting cell library; each plasmid comprises a connecting unit with mutually different base sequences; and Bsa I enzyme digestion sites are provided at junctions of the two tail ends of each connecting unit and an original carrier. The DNA library disclosed by the invention is adopted to construct transcription activator like effector nuclease plasmids; enzyme digestion and connection of the identifying modules can be carried out in one reaction, so that 12-19 identifying modules can be further connected, and therefore, the method is quick in speed, high in efficiency, simple to operate, easy for storing materials and low in cost.

Description

technical field [0001] The invention relates to the field of genetic engineering, in particular to a Tale junction unit library, a DNA library comprising all Tale junction units in the Tale junction unit library and a method for constructing a transcription activator-like effector nuclease plasmid. Background technique [0002] It has always been the dream of many scientists to modify the genome according to human wishes. Specifically delete or add the sequences we need on the endogenous genome. On the one hand, various animal models can be constructed for basic biological research and disease mechanism research. On the other hand, animal reactors can be produced to produce what we need cheaply. Biological components that are difficult to obtain from other sources. People have not found a simple and efficient method for genome-targeted modification of the genome. Traditional gene targeting technology relies on the random exchange of homologous chromosomes naturally occurri...

AI Technical Summary

Problems solved by technology

But they all have their own defects, because the chemical synthesis of highly repetitive sequences of DNA to be synthesized is very difficult and the cost is very high; the two-step method requires two steps of connection, so the material cost, time cost, and sequencing cost are all high; the now public The only method that can be connected in one step can only connect up to 14 recognition modules, and the length of common recognition modules in nature is 12-23. The limitation not only affects the application of this method, but also is not conducive to the improvement of TALEN specificity, and because of the need for enzyme digestion, purification, and re-ligation, the operation is cumbersome, the process is complicated, and the DNA after enzyme digestion and purification is difficult to store, especially single-stranded DNA. tail easily degrades

[0006] In the past, some researchers connected TALEN with a single module, such as connecting 18 recognition modules. It is very difficult to connect 18 fragments in one reaction, and no one in the world has been able to achieve it; Can connect 14 identification modules

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