Carya cathayensis auxin efflux carrier protein CcPILS gene cloning and expression analysis method

Abstract

The invention discloses a Carya cathayensis auxin efflux carrier protein CcPILS gene cloning and expression analysis method. The method includes the steps of: material preparation, RNA extraction and purification, CcPILS gene RACE, bioinformatics analysis, and temporal and spatial expression analysis. The method provided by the invention clones Carya cathayensis PIL homologous gene CcPILS by RACE technique, the gene has a full-length sequence of 1541bp, wherein the open reading frame sequence is 1263bp, bioinformatics software analysis is applied to predict that the sequence encodes 420 amino acids, the CcPILS protein molecular weight is about 46.22KD, PI is 5.38, the protein is located at an endoplasmic reticulum membrane, is equipped with 5 transmembrane hydrophobic structural domains respectively at the N terminal and C terminal, and is separated by a middle hydrophilic loop. The gene has high homology with Arabidopsis thaliana AtPILS5 and AtPILS7, and belongs to Clade III subtribe of PILS gene family. Fluorescence quantitative RT-PCR analysis shows that before and after Carya cathayensis grafting, CcPILS has expression trend in scion consistent with that in rootstock, and provides a beneficial promoting effect on solution of the problems of difficult Carya cathayensis grafting, difficult variety breeding and difficult quality improvement, etc.

Description

[0001] 【Technical field】 [0002] The invention relates to the technical field of pecans, in particular to a method for cloning and expression analysis of the pecan auxin export carrier protein CcPILS gene. [0003] 【Background technique】 [0004] Plants regulate auxin content in different tissues through auxin metabolism and intercellular transport, and then generate signals to regulate plant tropism, the occurrence and development of lateral organs, etc. Polar transport is an important feature of auxin, which is coordinated by the auxin input carrier AUX / LAX protein family (AUXIN RESISTANT1 / LI-KE AUX1) and the export carrier PIN-FORMED (PIN) protein family. Barbez et al. found another auxin intracellular transport-related gene family with 7 members. Although the sequence similarity between this gene and PIN is very low (10%-18%), its topological structure is similar to that of PIN protein, so it is named PILS (PIN-LIKES, PILS). PILS can be divided into three subfamilies, Cl...

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Problems solved by technology

[0007] The purpose of the present invention is to overcome the above-mentioned deficiencies in the prior art, and provide a method for cloning and expression analysis of the hickory auxin export carrier protein CcPILS, which aims to solve the problems of hickory grafting, variety selection, and quality in the prior art. raised technical issues

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