Method for constructing cotton bacterial artificial chromosome (BAC) library by non-dark cultured leaves

Abstract

The invention relates to the field of molecular biology and especially relates to a method for constructing a cotton bacterial artificial chromosome (BAC) library by non-dark cultured leaves. The method comprises the following steps of 1, selecting cotton leaves, 2, extracting a high-molecular weight nucleus DNA, 3, partly digesting the high-molecular weight nucleus DNA and carrying out selection twice, 4, carrying out recovery, connection and transformation of a large-fragment DNA, and 5, carrying out detection of an insert fragment. The method is suitable for construction of BAC libraries of all cotton materials which comprise cotton materials having low seed amounts and persistent cotton materials having no seed.

Description

technical field [0001] The invention relates to the field of molecular biology, in particular to a method for constructing a cotton BAC library using non-dark cultured leaves. Background technique [0002] At present, most of the constructed cotton Bacterial artificial chromosome (BAC) libraries are cultivars, and the selected materials are the cotyledons of etiolated seedlings obtained from the dark culture of cotton seeds after accelerated germination. The dark culture treatment can greatly reduce the production of chloroplast and phenolic substances, reduce its influence in the DNA extraction process and make the cells easy to rupture, and it is easier to obtain purer DNA from seedlings cultured in dark. However, the traditional cotton BAC library construction method has a narrow range of materials and requires dark culture treatment of the materials, which is complex and time-consuming; the construction of the BAC library requires a large amount of materials, generally m...

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Problems solved by technology

However, the traditional cotton BAC library construction method has a narrow range of materials and requires dark culture treatment of the materials, which is complex and time-consuming; the construction of the BAC library requires a large amount of materials, generally more than 40g. The traditional cotton BAC library construction method The amount of seeds used is large, and it is difficult to use materials with hard seed coats and difficult germination. It cannot be used for BAC library construction of cotton materials with few or no seeds.

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