Expression Of Three Lectin Genes (sGNA, SNTL, Pta) In Transgenic Wheat Lines Enhanced Resitance To Aphids

Wheat has a special status in stabilizing China food supply. The product loss of wheat due to insects is very high in recent years. For a long time the control of these pests has depended chiefly on large amounts of insecticides sprays which cause considerable environmental pollution and represent a health hazard to farmers as well as significantly increasing the costs of wheat production. Conventional plant breeding programs have been undertaken and considerable time and effort has been made in searching for insect-resistant germplasm resources, but it has moved slowly. Genetically modified (gm) can make up the deficiency of the conventional wheat breeding and breakthrough the limitation of available gene pool. Protease inhibitors, Bt endotoxins and plant lectins have been proven to be very successful in control insects. A more complete transformation system of particle bombardment method has been established, with young embryo and young ear as the receptor, which made it is possible to fostering high-quality and aphid-resistant wheat by genetic engineering.Plant lectins had been shown toxic to sucking pests resulting in reduced survival and delayed development. It’s significantly for developing aphid-resistant wheat germplasm resources by introducing these genes into common wheat.In this study, sGNA (synthesized Galanthus nivalis agglutinin) and sNTL (synthesized Narcissus tazetta lectin) genes encoding mannose binding lectins were synthesized according to the codons preferable principle of wheat genes and transgenic sGNA and sNTL common wheat lines, driven by constitutive and phloem-specific promoter rbcs, were generated by particle bombardment transformation. The objectives of this study were:(1) to confirm that the two synthetic genes sGNA and sNTL can be effectively expressed in transgenic wheat respectively;(2) to examine the aphid resistance performance of sGNA transgenic lines and sNTL transgenic lines indoors as well as field conditions and evaluate the functions of sGNA and sNTL against wheat aphid; and (3) to produce transgenic lines that may be useful for developing aphid-resistant wheat germplasm resources. Main research results are as follows:(1)42To sGNA transgenic plants,56To sNTL transgenic plants and54To pta transgenic plants were obtained by particle bombardment transformation with embryogenic calli as receptor. Twelve homozygous T4sGNA transgenic lines and eleven homozygous T4sNTL transgenic lines were identified by PCR amplification. The three genes were successfully integrated into the wheat genome and expressed stable according to the tests of PCR, RT-PCR、Southern blot. Northern blot and clotting test.(2) The transgene insert site in single gene patental line zy2-18containing sGNA were analyzed preliminary. The flanking sequences of sGNA were cloned by genomewalking, and located on chromosome A by the homologous recombination and random insertion.(3) After the insect bioassays in the lab and field-test, five sGNA transgenic lines (zy2-11、zy2-18、 jd8-10、yn5-2、yn5-20) and seven sNTL transgenic lines (fy4-4、fy4-10、af7-8、af7-10、zy9-1、 zy9-4、zy9-8) were found highly resistant to aphids compared to the corresponding control plants. Significant differences were also observed for mortality and growth rate of aphids between the transgenic lines and the corresponding controls in lab condition on detached leaf tissues. The aphid amount per tiller of af7-8sNTL transgenic line was reduced by80.3%maximum compared to the control Af9. Six pta transgenic plants showed a higher level of resistance genes in wheat□ccordingto the identification of detached leaf segment.(4) The sGNA transgenic lines and the controls showed a significant positive correlation (r=0.812*) between the relative aphid amount per tiller and OD values of leaf tissues extracts. The sNTL transgenic lines, of which it showed a significant positive correlation (r=0.932**) between aphid amount per tiller and OD values of leaf tissues extracts. So the result of clotting activity test indicated that the increased resistance to aphids was significantly correlated with the expression of sGNA and sNTL lectins in the transgenic lines.Therefore pta, sGNA and sNTL could be served as alternative choice to produce aphid-resistance wheat through genetically modified approach. It may also be worth commenting on the strategies for pyramiding the lectin genes.