Molecular biological method for identifying seedling stage of nicotine transformation tobacco plant

Abstract

The invention relates to a molecular biological method for identifying the seedling stage of a nicotine transformation tobacco plant. The method comprises the following main steps of: extracting the genomic DNA from a fresh leave sample of the plant in the seedling stage by a CTAB (hexadecyltrimethylammonium bromide) method, performing PCR (polymerase chain reaction) using the genomic DNA as an amplification template in combination with specific primers, then enabling the PCR products to be subjected to agarose gel electrophoresis, observing and reading a molecular marker spectrum in the gel imaging system, and subsequently comparing the molecular marker spectrum with the target spectrum to determine the transformation characteristics of the sample. The method in the invention is free from environmental factor influences, is in no need of artificial chemistry to induce nicotine to transform, and is capable of sampling operation during the seedling stage to bring forward the identification and screening of a transformation plant ahead of tobacco plant implantation, thus greatly saving time, being accurate and efficient, simple and convenient to operate, easy to popularize, and applicable to purification of tobacco parent seeds and identification of nicotine transformation traits as well as identification and removal of nicotine transformation plants of other tobacco types.

Description

technical field [0001] The invention relates to an identification method for tobacco nicotine transformed strains, in particular to a molecular biological identification method for tobacco nicotine transformed strains at seedling stage. Background technique [0002] The current research results show that there are four main types of alkaloids in tobacco: nicotine, nornicotine, anatabine, and anatabine. Common tobacco belongs to the nicotine accumulation type, mainly nicotine, its content accounts for more than 90% of the total alkaloids, nornicotine is the second largest alkaloid, the content generally only accounts for 3-5% of the total alkaloids, the other two Alkaloids account for only a small part. The level of nicotine content is an important factor to determine the sensory quality of tobacco leaves. Generally, it is required that the content of nicotine should be appropriate to achieve the purpose of moderate physiological strength, mellow taste and low irritation. N...

AI Technical Summary

Problems solved by technology

Although this method is effective, there are following deficiencies: 1, the sampling period is still late in the group tree stage after transplanting, and the field was wasted after removing the transformed strains, and it is difficult to ensure the size of the target group; 2, it needs to be induced by chemicals. ;3. The identification results are affected by environmental factors: such as drug concentration, modulation temperature and time, chemical detection factors, etc.

Therefore, although there is a good reference, it is still not efficient and timely enough to meet the actual needs