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Cloning and Analysis of CONSTANS-like Genes in Wheat (Triticun Aestivum L.)

Author: SuYaZhong
Tutor: LiSuoPing; JiaJiZeng
School: Henan University
Course: Genetics
Keywords: Wheat (Triticun aestivum L.) CO-like gene Genetic mapping Functional verification
CLC: S512.1
Type: Master's thesis
Year: 2012
Downloads: 56
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Abstract


Photoperiodism is an important physiological phenomena in plants which controls the flowering timeby perceived the length of day. Photoperiod has greatly effects on plant growth period, adaptability andyield. CO (CONSTANS) is an important gene in plant photoperiodic pathway, in recent years the study ofCO gene family has made a great progress in the model species, such as rice and Arabidopsis, but it is lessin wheat, which is the main food crop. So that cloning and analysis the structure and function of CO genesin Wheat has the important theoretical and practical significance.This experiment had cloned three CONSTANS-like genes (TaCOLX, TaCOL4and TaCOL5) in wheatby homology cloning and cDNA library screening; researched their structures, homology and sequencepolymorphism by sequence analysis; and transform rice through agrobacterium-mediated technique todiscuss the function of TaCOLX and TaCOL4. The main progresses were showed as follows:1. Three CONSTANS-like gene family genes (TaCOLX, TaCOL4and TaCOL5) had been cloned byhomology cloning and cDNA library screening. TaCOLX gene sequence have two exons and one intron, itsCDS sequence has an ORF which is960bp lengh and codes320amino acids, it also contains two B-boxdomain and one CCT domain; TaCOL4gene sequence has one exons, its CDS sequence have an ORFwhich is1266bp lengh and codes422amino acids, it also contains one B-box domain and one CCTdomain; the CDS sequence of TaCOL5gene has an ORF which is978bp lengh and codes326amino acids,it also contains one B-box domain and one CCT domain; B-box domain and CCT domain are the typicaldomain of CONSTANS-like gene family, that is to say each of TaCOLX, TaCOL4and TaCOL5can code aCONSTANS-like protein.2. Designing the nonspecific primer of A, B and D genome for TaCOLX and TaCOL4gene, thenamplifyed the wheat of Chinese spring and got the three homologous genes of TaCOLX which named asTaCOLX-A, TaCOLX-B and TaCOLX-D, their similarity of nucleotide sequence is more than97.1%; thesame way I have got the three homologous genes of TaCOL4which named as TaCOL4-A, TaCOL4-B andTaCOL4-D, and their similarity of amino acid sequence is more than97.6%.3. To analyse the sequence polymorphism of TaCOLX gene and TaCOL4-D gene in eleven parent wheats. Discovered that TaCOLX-A have one deletion of9bp lengh in the5’UTR region, TaCOL4-D havetwo SNP locus in coding region and fifteen SNP locus outside of the5’UTR region in the parent wheats.4. To locate the TaCOLX-D gene and TaCOL4-D gene by using the Chinese Spring nulli-tetrasomiclines. The TaCOLX-D was located on wheat chromosome2D; designing a mark through the different ofTaCOL4-D gene’s an alleles between M6and Opata85, and use the mapping group of M6×Opata85, theTaCOL4-D was mapped between maker Xfba8and Xgwm111of wheat chromosome7D with the geneticdistances of4.0cM and1.7cM.5. Building the vector of TaCOLX gene and TaCOL4gene to transform rice. And have separately gotthe T0generation transgene rice of TaCOLX and TaCOL4which generation is delaying heading. Toidentify their T1generation transgene rices and discovered that the generation of delaying headingseparated with the genotype, so TaCOLX and TaCOL4delayed the heading of rice.

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