Construction Of The CDNA Library Of Leymus Chinensis And Cloning And Analysis Its Photosynthesis Genes In The Grassland Reconstruction

At present, the main problems with ecological environment in China are overgrazing, grassland degradation, worsened desertification, loss of soil and water, reduction of species and biodiversity and severe ecological environment, which all restrict social sustainable and healthy development of social economy. The aim of the vegetation rehabilitation is vegetation construction. The vegetation rehabilition should be built artificially, the type of which should be made suitable for the present eco-environment.Vegetation construction must be in line with the fundamental principle of balanced ecological rationality, efficiency and sociability. With this principle, selecting and cultivating grazing variety capable of adapting to impoverished environment conditions, high value and ecology efficiency is a very important research project and assignment. The goal of the present research is using the molecular biological technique to cultivate the superior variety.Leymus chinensis belongs to Gramineae Leymus, it is a type of perennial rhizome forage, which has many advantages such as ample protein, high productivity, good palatability, extensive adaptation, and high tolerance to drought, salt and alkaline. Leymus chinensis plays an important role in the development of the livestock husbandry, grasslands restoration and desertificatiom controlling. Recently, problems like land desertification, overgrazing and the heredity feature of Leymus chinensis, such as low seed setting rate, low seedling emergence rate and low productivity, all the factors have very negative effect on predominance of Leymus chinensis, and severely hindered the sustainable development of building artificial grasslands, improving natural meadow and constructing ecological environment. The most urgent research project now becomes a fundamental research on the biology of Leymus chinensis, since most available reports from home and abroad focus on the macro field of grassland ecology and there is scarcely any systematic study on the biology of Leymus chinensis. In order to improve productivity of Leymus chinensis, we report the construction of the leaf of Leymus chinensis cDNA library. We used the M13 universal primers to screen positive clone to sequence and got a Rubisco large subunit gene and a gene fragment of Rubisco small subunit, the amino acid sequence had a high homology compared with other species. The main results obtained were following:1. A cDNA library was constructed with SMART cDNA Library Construction Kit using the RNA from the leaf of Leymus chinensis. The primary library has a high titer of 106cfu/ml and the amplified library has a titer of 1011cfu/ml, in which 98% clones are recombinant and the length of insert cDNAs is from 0.5 kb to 2.5 kb. The positive signals of TC and TMT of vitamin E in Leymus chinensis are detected by PCR in the amplified library. This high quality cDNA library provides a useful tool to study the molecular mechanisms of the Leymus chinensis.2. Base on the leaf cDNA library of Leymus chinensis, we used the M13 universal primers to screen positive clone to sequence. The sequence was submitted to NCBI (National Center for Biotechnology Information) for estimating its homogeneity (Genbank accession: Leymus chinensis CV Jisheng No.1 leaf cDNA library).3. By blasting with database of Genbank, we got a Rubisco large subunit gene and a gene fragment of Rubisco small subunit, the amino acid sequence had a high homology compared with other species. Rubisco large subunit gene was 1796 long, which was more than 98% homologous to the sequences of Rubisco large subunit gene nucleotides in Hordeum capense,Triticum aestivum,Bromus arvensis,Aegilops tauschii,Eremopyrum triticeum,Heteranthelium piliferum et al. ORF of Rubisco small subunit gene was 186bp long and encoded 61 amino acids. It was 93%,93%,91%,91% and 92% homologous to the sequences of amino acid in Hordeum capense,Triticum aestivum,avena clauda ,secale cereale and bromus cathartic,respectively. The cloning of Rubisco genes is useful for the further research of photosynthetic efficiency in Leymus chinensis.