Cloning Of Candidate Gene To Spike Differentiation In Maize

The kerenal number per ear is consisit of raw number and kermel per row in maize. In the phenotype of the two important component factors, it exhibited typical quantitative characteristics, and were formed in the developmental procedure of the spikelet and floret differentiation, controlled by a set of qualitive genes in seed development. Moreover, the mutant these genes would lead to low fecundity. In the female spikel and male spikel formation procedure, a series of transformation and differentiation in cell and organ were processed: from vegetative growth to reproductive growth, germ cells to the spikelet pair meristem, spikelet pair meristem to spikelet meristem, spikelet meristem to floret meristemand. Out of the genes exhibited in the whole developmental process, whichever occuring mutantion might lead to abnormal phenotype of maize spikelet or floret, and decreased the seed setting in maize. So, cloning of the candidate genes related to spike and floret differentiation in maize, not only can clarify the genetic mechanism of the formation of maize ear, but also can find agvangeous genes controlling the ear fecundity of maize. In this study, the mutants related to maize ear differentiation were selected in the field by using transposon tagging, and its flanking sequnence were cloned by means of a kind of modified AFLP method. The main results were showed as fellow:1. Using the method of Mu transposon insertion,four mutants of the ear differentiation were found in a 29,732 F1 progeny plants between inbred line 87-1(Rf4Rf4) and Mu7,it expressed male sterility, decreased the number of tassel spikelets, and reduced the number of lower solid(resulted 4-10 seeds per panicle)。2. A special amplification band between two parents and the mutants was selected using a modified approach named Mu-AFLP method, and an about 800bp AFLP band was found when using a nested Mu7-specific primer BglⅡ-2 and a selective primer M1. After sequenced the specific amplificated fragment, two different gene fragement contained partial sequences of Mu transposon and two unknown fragments were found. One gene fragement including two 220bp, had 88% similarity with gene rsp4, which related to cell differentiation; and the second candidate gene fragement had 99% similarity with NADH dehydroqenase subunit 1, which was found BAC AY928077.1 of B73.3. Using the method of electronic splicing in Genbank, the cDNA of frist candidate gene obtained a 509bp, and had 68% similarity with gene rsp4, the full length of genomic DNA in the B73 sequence database was 1194 bp. The candidate gene 2 was consisted of 2432 bp genomic DNA based on B73 sequence database. The two candidate genes whould be determined by using real time-PCR and transgenic method.