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Effects of Cs-ACS1 Transgene on Sex Differentiation of Cucumber

Author: QuShuJuan
Tutor: ZhangXingGuo
School: Southwestern University
Course: Olericulture
Keywords: Cucumber Cs-ACS1 gene Sense expression vector Antisense expression vector Transgenic Sex differentiation
CLC: S642.2
Type: Master's thesis
Year: 2010
Downloads: 47
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Abstract

Breeding full estrogen or strong female cucumber varieties is one of the main objectives of the cucumber breeding. Cucumber female flower node appears early, and number are distributed, cucumber early maturing, high-yielding, stable yield basis. Gender differentiation has clearly cucumber addition to the influence of the master genes, but also by a variety of factors, endogenous ethylene is one of the important factors. Many researchers believe that the cucumber ACC synthase gene control female F locus tightly linked. Where, Cs-ACS1 gene and F gene had high homology the early F gene mistaken. Visible, cucumber Cs-ACS1 gene and cucumber female differentiation there is a correlation. In order to verify the Cs-ACS1 gene transfected gene cucumber sex differentiation, this test uses a PCR method from the full estrogen is cucumber 0401 genomic DNA was amplified out 2333 BP's gene fragment, build into justice, antisense gene expression vector as well as by the gene promoter to control the expression of red fluorescent protein reporter gene carrier, to import cucumber, and the purpose of the three plant expression vector gene analysis confirmed Cs-ACS1 gene with cucumber female sex differentiation is closely related to observation of transgenic plants. The main contents of the test and research results are as follows: 1. Cloned Cs-ACS1 gene cloning Cs-ACS1 gene by BLAST analysis showed only one base difference: the cloned gene sequence Cs-ACS1 published , the nucleotide sequence homology of 98%, and the deduced 247 amino acids is completely consistent. The above results show that: the cloned gene for Cs-ACS1 gene. And justice genes of 2.Cs-ACS1 import cucumber co-suppressed transgenic cucumber plants male of the Cs-ACS1 gene the insert carrier pVCT2102 build just the gene plant expression vector pVCT2214. After digestion and PCR detection using Agrobacterium-mediated method the Cs-ACSl justice genes into strong female lines cucumber 0837 (the laboratory breeding varieties). The after transplanting of 12 survived transferred gene cucumber plants PCR detection preliminary determine Cs-ACS1 justice genes have been transferred to the cucumber genome; survival of plant phenotype observed, found that 12 transferred genes cucumber plants 10 all open male flower, All plants the terminal bud is the male flowers that Cs-ACS1 gene with strong female cucumber body of Cs-ACSl gene mutual inhibition, limit itself Cs-ACS1 gene expression, as well as restrictions transferred to the Cs-ACS1 gene expression, that appeared in a total of inhibition. Antisense gene in 3.Cs-ACS1 transgenic cucumber plants can cause virilization justice Plant expression of the carrier pVCT2214 by restriction enzyme digestion, antisense plants expression vector pVCT2215. Been tested to determine the vector was successfully constructed using Agrobacterium-mediated method the Cs-ACSl antisense gene into a strong female cucumber 0837. After transplanting, the turn genes cucumber Total survived two on the survival of transgenic cucumber detection (PCR), to determine Cs-ACS1 antisense gene has been transferred to the cucumber genome; observe survived two anti-justice turn genes cucumber, found that plants were all open male flower , and all plants the terminal bud male flowers, speculated that antisense inhibition. 4.Cs-ACS1-REF gene can be expressed in the flower buds of transgenic cucumber Cs-ACS1 gene was inserted into a vector containing a red fluorescent protein, and the resulting plant expression vector pVCT2217. Infection with Agrobacterium EHA105/pVCT2217 broth cucumber (strong female 0837) cotyledon six cucumber plants survived the turn genes Cs-ACS1-REF, culture, all open male flower. Take turn Cs-ACS1-REF gene cucumber genomic DNA by PCR, and determine the transfer CsACS1-REF genes have been transferred to the cucumber genome. Whitening treatment a week after, fluorescence microscopy detected to turn Cs-ACS1-REF cucumber flower buds red fluorescent groups that Cs-ACS1-REF gene expression in plants in flower buds.

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