Sequence Analysis Of α-gliadin Gene 5' Upstream Regulation Region And Coding Region In Triticum Compactum Host.

According to the conserved sequences ofα-gliadin gene 5' upstream regulation region and coding region,the gene-specific primers were designed to amplify the 5' upstream regulation region and coding region from Triticum compactum Host.The PCR amplyfied fragments were sequenced,and investigated.The results were as follows:1.Fourty-oneα-gliadin gene coding region sequences,included 11 sequences from PI566594,11 sequences from PI269250,4 sequences from PI191874,3 sequences from PI159101,12 sequences from PI352302,were obtained.The structure characters ofα-gliadins gene were identified in all sequences,including the singal peptide,repetitive domain,polyglutamine regionⅠ,unique regionⅠ,polyglutamine regionⅡ,unique regionⅡand the toxic sequences(PSQQQP).There were 25 sequences which had in-frame stop condons,and the main type of in-frame stop condons was mutation of CAG→TAG. N-terminal,included 5 amino acid residues,was confirmed and separated from the repetitive domain.The repetitive domain was deduced the same repetitive motifs PQPQPFP,PQQPY and LPYPQPQ,which difference was that variable number of repetitive motif LPYPQPQ was involved in different sequences.At the two polyglutamine regions,the extensive different of length among all sequences were found,and some other amino acid residues were also detected,besides glutamines.The mutations were mostly happened at the first base in codons.Four cysteine residues were detected in sequences PI269250-1-C,PI566594-9-B and PI566594-2-A,and six residues in sequences PI269250-6-B,PI269250-8-C,PI269250-11-C,PI566594-5-A,PI566594-7-B and PI352302-5-C,but five cysteine residues including four in unique-Ⅰregions and one in unique-Ⅱregions were detected in the other sequences.The dendrogram were constructed based on the nucleotide sequences of the coding domain and amino acid sequences.The clustering results were consistent except sequences PI352302-2-C,PI269250-9-C, PI566594-2-A,PI159101-1-A,PI159101-2-A and PI191874-3-C.Using the previous 23 sequences,which chromosome sites were already known,these obtained sequences could be located to special chromosome.The result based on the nucleotide sequences of the coding domain(deleted repetitive and polyglutamine domains) suggested the sequences PI269250-2-C,PI566594-11-C,PI352302-11-D,PI352302-3-C,PI269250-7-B,PI352302-6-C,PI566594-6-B,PI352302-1-B,PI352302-4-C,PI269250-1-C,PI352302-10-B,PI191874-4-C,PI352302-12-D,PI191874-1-B,PI352302-7-D,PI566594-8-B,PI566594-I-A,PI269250-5-B,PI566594-9-B,PI566594-10-B,PI269250-8-C,PI566594-5-A,PI269250-11-C,PI566594-4-A and PI269250-3-C were closely related to the genes on chromosome A,sequences PI566594-3-A,PI269250-10-C,PI159101-3-B,PI352302-9-B,PI352302-5-C,PI566594-7-B,PI269250-6-B,PI352302-8-B,PI566594-2-A,PI159101-1-A,PI159101-2-A and PI191974-3-C were closely related to the genes on chromosome D,and sequences PI269250-4-D and PI269250-9-C were closely related to the genes on chromosome B.2.Fifty-oneα-gliadin gene 5' upstream regulation region sequences were obtained.The length of all sequence varied from 216bp to 436bp.Nineteen sequences,whose length exceeded 380bp,were analyzed.Sequencing analysis showed that the number of adenine was first,that of thymine and cytosine were second,and that of guanine was third.The average homology with the 19 sequences was 92%,the lowest similarity was 83%,and the highest was 100%.Compared with other species,the average homology was 71.9%.These sequences had closest homology with sequence X54688 from T.aestivum(92.9%),higher homology with the sequence from Dasypyrum breviaristatum(91.8%),and lowest homology with sequence AY998010 from H.chilense(46.0%).Fragrnent(51bp) was absent in sequences PI566594-I5-G,PI566594-16-G,PI269250-1-C,PI191874-2-C, PI191874-3-C,PI159101-5-G and PI352302-2-C.And the 19 sequences all included important regulatory elements associated with gene expression.For example,Endosperm Motif(EM),GCN4-Like Motif,CAAT-Iike box,AAAG Motif,TATA box,CAC-box and AACA motif.The sequences aligument indicated that there had rich variation inα-gliadin gene 5' upstream regulation region of T.compactum,T.aestivum and T.urartu.The clustering results indicated the genetic relationship in T.compactum and other species based on 5' upstream regulation region was complex.According to the clustering results,it could not distinguish the material resource,and also could not find the specific site on chromosome.