Flanking Sequences Of Maize DNA In The Study Of Rice Mutant

The aerial roots of maize mutant lines of rice traits which has eight generations of genetic stability was obtained from 2000 via distant molecule hybridization created in our laboratory. In this thesis, the variatied materials which inherited steadily with maize characteristics included aerial root, were selected for study and they were numbered 08-7(+2)-131-5-2(91),CK1(Zhengdan 14),and CK2(Xiangdao 1)respectly and they were analyzed through the molecular maker technology of AFLP for polymorphism, there was one band was examined between variant line and positive control, called the target band which was cloned and sequenced and the length of the target band is 349bp The Blast searchs the result, the sequence with100% similarity of the maize sequence (ref | NM₀01155903.1). Nested primer P5M3 Sense and Anti Sense-P5M3 were selceted according to maize sequence for detecting between positive control and Variants Lines of validation, a 212bp band geted via gel extraction and sequenced, the positive control and variation of the same strains was 99.53%, with only one base difference after DNAman analysis the Blast search further determine the sequence of maize sequence.TAIL-PCR technique using the known target sequence of unknown flanking sequences extended, According to the target band of DNA of known sequence were designed for three specific nested primers SP1; SP2; SP3, according to the known DNA sequence of the corresponding protein (NCBI search cysteine disulfide bonds on Blastp enzyme / ubiquitin Protein, conserved protein sequences based on degenerate primers) sequence designed five degenerate primers that Tail-AD1; Tail-AD2; Tail-AD3; Tail-AD4; Tail-AD5, via three specific primers and the five degenerate primers in any one combination, by optimizing the primer combination and reaction system, in which degenerate primers Tail-AD3 and specific primer combination is the ideal result. Obtained a long DNA sequence to 2123bp by sequencing and further retrieve the sequence in MaizeGDB, the results showed that the sequence locates in chromosome 2 of maize, the match rate was 94%, NCBI Blastp search revealed the sequence of the maize mRNA NM₀01155903 sequence in (Encoding cysteine disulfide enzyme/ubiquitin-protein), the biological software- DNAman analyze the DNA sequence encoding a 449 amino acid protein sequence, named h-cys protein, the sequence NP₀01149375 the consistency was 81%. Sequence variation of rice that contains a large fragment of maize DNA sequences, we concluded that the variation of materials and the insertion of maize or recombinant DNA certain degree of relationship, further proof of sub-hybrid rice is far fate of maize DNA fragments can cause rice cells mutate .