In-vitro identification method for gramineal self-incompatibility phenotype

Abstract

The invention belongs to the technical field of plant biology, and discloses an in-vitro identification method for gramineal self-incompatibility phenotype, which comprises the following steps: (1) inflorescence arrangement: the inflorescences of a paternal plant are pruned at the full-bloom stage, removing bloomed spikelets; (2) artificial flower forcing: the inflorescences are inserted into Hoagland nutrient solution, and flower forcing treatment is carried out under a completely dark condition for 8 to 12 hours; (3) in-vitro pistil culture: mature spikelets are chosen from the maternal inflorescences, and the pistils are intactly cut off under a stereoscopic microscope and inserted into 0.5 to 1 percent of agarose solid medium for in-vitro culture; (4) indoor pollination: in the morningof the next day, paternal pollen is used for pollinating the pistils; (5) pistil softening: the pollinated pistils are softened with 8 to 10mol / L of NaOH for 3 to 4 hours; (6) pistil dyeing: the softened pistils are dyed with 0.1 to 0.5 percent of aniline blue; (7) microscopic observation: the growth of the pollen tubes on the pistil stigmas are observed in an inverted fluorescence microscope. The method is simple and easy to master, the accuracy is high, and the method is suitable for being applied in the genetic analysis and crossbreeding research of miscanthus.

Description

Technical field [0001] The invention belongs to the field of plant biotechnology, and specifically relates to an in vitro identification method for the self-incompatibility phenotype of the Gramineae. Background technique [0002] Miscanthus (Miscanthus) is an important class of C4 gramineous energy grass and forage grass. It has the characteristics of high biomass production and strong stress resistance. It is naturally distributed in East Asia and Southeast Asia. my country mainly has awn (M.sinensis), M. floridulus (M. floridulus), Di (M. sacchariflorus), South Di (M. lutarioriparius) and other species, they all have strict self-incompatibility (Self-incompatibility, SI), that is, self-incompatibility. Self-incompatibility is of great significance for plants to avoid inbreeding and maintain genetic diversity. It is also one of the main ways to use plant heterosis. Self-incompatibility is used as the female parent to make it compatible with other self-incompatibility. Crossing ...

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Problems solved by technology

[0003] But on the other hand, self-incompatibility plants have been sexually reproduced by hybridization in nature for a long time, and the heterozygosity of individual genomes is high, and the genetic background is complex. Since it is impossible to obtain self-bred pure lines, traits segregate in the hybrid F1 generation , not only reduces the performance of its heterosis, it is not suitable for large-scale production of hybrid seeds to promote planting, but also greatly limits the development of genetic research such as gene mapping, gene map cloning and whole genome sequencing, hindering further genetic improvement, Therefore, it is necessary to find a way to overcome its self-incompatibility

[0004] In the hybrid breeding and genetic research of Miscanthus and other plants, it is often necessary to measure the self-incompatibility index or the hybrid compatibility index between different genotypes. At present, the self-incompatibility index or the hybrid compatibility index The conventional measurement method of self-incompatibility phenotypes such as index is to use artificial bagging to force self-pollination or artificial cross-pollination on field or potted plants. After the plants enter the mature stage, the self-incompatibility or For the seed setting rate of hybridization, the test cycle of this method generally takes several months, which is not only time-consuming and heavy workload, but also the test results are easily affected by external environmental conditions such as temperature, humidity, wind force, rainfall, etc., resulting in poor repeatability of the test , the accuracy is not high

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