Purification method of single-stranded binding protein and application of single-strand binding protein in gene synthesis

Abstract

The invention relates to a purification method of single-stranded binding protein, an application of the single-stranded binding protein in gene synthesis, and a sequential assembly method of a groupof single-stranded or double-stranded DNAs with homologous sequences. The method is applied to assembly of single-stranded nucleic acid molecules with homologous sequences, the single-stranded nucleicacid molecules serve as an additive in a polymerase chain reaction, and the method is applied to assembly of double-stranded DNA molecules with homologous sequences. The specific scheme comprises plasmid cloning of the high-temperature-resistant SSB protein, protein purification and an application in gene synthesis. By applying the ssb protein, the splicing assembly of the single-stranded or double-stranded DNAs is accelerated, and the assembly accuracy is improved.

Description

technical field [0001] The invention relates to the technical field of assembly of nucleic acid molecules, in particular to a method for purifying a single-chain binding protein and its application in gene synthesis. Background technique [0002] Synthetic biology is an important field in molecular biology. Gene synthesis includes the synthesis of primers or double-stranded DNA fragments as short as tens of bases, as well as the synthesis of genes ranging from tens of thousands to hundreds of thousands of bases. [0003] The basic method of gene synthesis includes two steps. In the first step, multiple chemically synthesized primers are mixed, annealed through the homologous sequences between the primers, the primers that annealed successfully are extended after adding DNA polymerase, and multiple PCR cycles Finally, the short primers form longer DNA fragments. In the second step, the desired full-length gene is amplified by adding primers at both ends to the product of the...

AI Technical Summary

Problems solved by technology

In the actual annealing process, primers are often annealed incorrectly. In addition to the correct primer annealing process, there are also two primers that should not be annealed in the reaction that anneal incorrectly, or anneal at the wrong position, or even anneal to the primer itself. These conditions reduce the efficiency of gene synthesis

Especially for genes with high AT and high GC, due to the high homology of the gene itself, the probability of misannealing of primers is also higher, so it is difficult to synthesize such difficult genes by means of PCR

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