【摘要】：大麥(Hordeum vulgare L.)屬禾本科植物,主要用于制造啤酒及飼用。其具有抗旱、耐鹽堿等特性。隨著全球氣候與生態(tài)環(huán)境的日益惡化,土壤鹽堿化和干旱仍然是限制大麥生長(zhǎng)的兩個(gè)主要環(huán)境因素。采用基因工程手段,培育高抗旱耐鹽性大麥新品種已成為大麥栽培的迫切要求。而受體材料是遺傳轉(zhuǎn)化的基礎(chǔ),尋求一種來(lái)源廣泛、取材方便、受基因型和季節(jié)限制小、高頻再生的體系對(duì)于有效地開(kāi)展大麥轉(zhuǎn)基因研究具有重要意義。因此,本實(shí)驗(yàn)通過(guò)建立大麥莖尖再生體系,利用農(nóng)桿菌介導(dǎo)法將鹽生草(Halogeton glomeratus)Na~~+/H~~+逆向轉(zhuǎn)運(yùn)蛋白(NHX)HgNHX1基因?qū)氪篼?獲得轉(zhuǎn)基因陽(yáng)性植株并對(duì)轉(zhuǎn)基因大麥T1代植株進(jìn)行抗逆性分析。主要研究結(jié)果如下:1.以栽培中常用品種甘啤3號(hào)、甘啤4號(hào)和甘啤6號(hào)為材料,研究了大麥莖尖高頻再生體系影響因素,其中種子浸泡在30%的硫酸中,30℃、180 r/min搖動(dòng)1 h為去除穎殼最佳方法;75%的乙醇處理1.5 min~~+2%的NaClO處理15 min為較好的滅菌方法;3 d苗齡、莖基部2 mm處切段效果最好;3 mg/L 2,4-D有利于愈傷組織誘導(dǎo),參試品種中甘啤4號(hào)再生能力較強(qiáng)。2.利用鹽生草HgNHX1對(duì)大麥進(jìn)行遺傳轉(zhuǎn)化,采用農(nóng)桿菌介導(dǎo)法,真空滲透處理5 min,超聲波處理30 s,100 r/min搖動(dòng)侵染30 min的瞬時(shí)轉(zhuǎn)化率較高,獲得了抗性再生植株23株,PCR檢測(cè)陽(yáng)性植株數(shù)7株,并獲得了T1代種子。3.對(duì)T1代轉(zhuǎn)基因大麥植株抗逆性分析發(fā)現(xiàn),在干旱、鹽脅迫下,HgNHX1基因在大麥中表達(dá)豐度增高,轉(zhuǎn)HgNHX1基因大麥植株的耐鹽性不明顯,而抗旱性增強(qiáng)。
[Abstract]:Barley (Hordeum vulgare L.) Gramineae, mainly used in beer and feed. It has the characteristics of drought resistance, salt and alkali tolerance and so on. With the deterioration of global climate and ecological environment, soil salinization and drought are still the two main environmental factors that limit the growth of barley. It has become an urgent requirement for barley cultivation to cultivate new barley varieties with high drought resistance and salt tolerance by means of genetic engineering. However, the receptor material is the basis of genetic transformation, so it is of great significance to find a system with wide source, convenient collection, small genotypic and seasonal restriction and high frequency regeneration for the effective research of barley transgene. Therefore, by establishing the stem tip regeneration system of barley, the (Halogeton glomeratus) Na~~ / H / H reverse transporter (NHX) HgNHX1 gene of halophytic grass was introduced into barley by Agrobacterium tumefaciens mediated method. Transgenic positive plants were obtained and stress resistance of transgenic barley T1 plants was analyzed. The main results are as follows: 1. The influencing factors of high frequency regeneration system of barley stem tip were studied by using the commonly used varieties Ganbei 3, Ganbei 4 and Ganbei 6 as materials. The seeds were soaked in 30% sulfuric acid at 30 鈩，

本文編號(hào)：2476423