Expression Analysis Of Cell Wall Invertase Gene TaCwi-Al And Cloning Of Dense And Erect Panicle1Gene And Development Of Functional Markers In Common Wheat

Improvment of grain yield is an important objective in wheat breeding, and development andapplication of functional markers of genes associated with grain yield is of great importance in wheatbreeding programs. Functional markers (FM) are developed from sequence polymorphisms present inallelic variants of a functional gene at a locus. FMs accurately discriminate alleles of a targeting gene,and are ideal molecular markers for marker-assisted selection in wheat breeding. To date, more than30wheat loci associated with processing quality, agronomic traits, and disease resistance, have been cloned,and97FMs were developed to identify93alleles based on the sequences of those genes.In the present study, the transcript expression pattern of cell wall invertase (CWI) gene TaCwi-A1associated with thousand kernel weight was analyzed during grain development in common wheat. Therice dense and erect panicle1(DEP1) gene OsDep1that controls panicle architecture was used as acandidate to clone wheat DEP1gene (TaDep1). The TaDep1was cloned and characterized, and itsallele-specific markers were developed. The main results obtained in this study are summarized below.1. Expression patterns of TaCwi-A1a and TaCwi-A1b were determined by semi-quantitativeRT-PCR with4TaCwi-A1a and3TaCwi-A1b genotypes at7,14,21,28and35dpa. During graindevelopment, putative transcription level of TaCwi-A1a and TaCwi-A1b alleles showed the highestexpression levels at the early developmental stage (7dpa), then decreased rapidly from7dpa to28dpa,and there were no expression at nearly mature stage (35dpa). TaCwi-A1a allele still expressed at28dpa,whereas no expression was detected at TaCwi-A1b genotype at28dpa, which may be a reason resultingin the differences of thousand kernel weight between two genotypes.2. Full-length genomic DNA sequences of TaDep1on the homoeologous group5chromosomeswere cloned from common wheat, which have five exons and four introns similar to that of rice OsDep1.The open reading frame (ORF) of TaDep1-A1, TaDep1-B1and TaDep1-D1were918bp,888bp and900bp, encoding polypepetides of305,295and299amino acids, respectively. The TaDep1-B1ofcommon wheat has high similarity with barley and Triticum urartu Dep1genes, with89.9%and93.3%of identity in deduced amino acids, respectively. The deduced amino acids of TaDep1-B1showed46.9%of identity with those of rice OsDep1.Five allelic variants were identified at TaDep1-A1locus, with11SNPs and1InDel. Allelic variantsTaDep1-A1a and TaDep1-A1b encode the same deduced amino acids, with DNA sequencepolymorphisms in the introns. The allelic variants of TaDep1-A1c, TaDep1-A1d and TaDep1-A1e alsohave the same deduced amino acids. There were13SNPs and4InDels among four allelic variants ofTaDep1-B1locus, and a30-bp deletion was present in the fifth exon of TaDep1-B1c. The deducedamino acids encoded by TaDep1-B1a and TaDep1-B1b were identical. TaDep1-D1locus has two allelicvariants, with2SNPs and2InDels. One amino acid difference was found between the deduced aminoacids encoded by TaDep1-D1a and TaDep1-D1b.Three pairs of complementary dominant markers and one co-dominant marker were developedbased on the sequence polymorphisms present in allelic variants of TaDep1-A1and TaDep1-B1. Four populations and over430wheat cultivars were used to validate the association of these markers withplant height, panicle length, spikelet number and thousand kernel weight No association was foundbetween the allelic variants of TaDep1-A1and TaDep1-B1and phenotypic data of yield, indicating thatthese genes have no effect on yield-related traits in common wheat. The co-dominant marker dep19,which can accurately discriminate the allelic variants TaDep1-B1c and TaDep1-B1c, TaDep1-B1b,TaDep1-B1d, was developed from a30-bp InDel of different allelic variants at the fifth exon ofTaDep1-B1.