Detection Method Research Of Maize Transgenic And Cytoplasmic Male Sterile(CMS)

Maize is an important food crop and feed crop.Molecular markers have been widely applied to corn authenticity,purity,detection of transgenic and cytoplasmic sterility.With the country’s concern for corn genetic safety,the detection of corn genetically modified genes is increasingly important.Maize male sterile seed production is gradually replacing conventional seed production methods.When male sterile seed production,two issues need to be solved.About Google Translate Privacy & Terms.First,before the male sterility is seeded,the purity of the female parent must be tested;the second is to test the purity of hybrids after male sterility.Based on the detection requirements of maize transgenic and cytoplasmic sterility seed production,a multiplex PCR detection system for maize transgenic and cytoplasmic sterility was established based on a fluorescent capillary platform.In the early stage,primers were designed for the 10 exogenous genes,zss11 b,Bt,PAT,NOS,Ca MV356 S,NPTII,PMI,HPT,CP4-1,and CP4-2.five layers of PCR were successfully constructed using five primers: maize zss11 b,BT,PAT,NOS,and Ca MV35 S.By adding G at the5’ downstream of the primer,the problem of the N+1 peak was solved.The number of PCR cycles in this system was determined to be 35 cycles;Through the adjustment of the concentration between primers,the sensitivity of the multiplex PCR detection system was improved,and the detection limit was less than 0.1%.In this study,using 29 representative maize inbred lines with rich phenotypes and genotypes and triplets materials,these included 21 inbred lines representing different heterotic groups,5cytoplasmic male sterility(CMS)materials,2 C-type CMS materials,and 1 T-CMS sterile material.Three sets of triad materials Jingke 968,Zhengdan 958 and Xianyu 335 hybrids and their respective parents.The primers of 11 InDel loci from chloroplast genome were designed,screened and evaluated,and two specific pairs primers to identify the maize S-type CMS were obtained.The variations of the two primers CPMIDP01 CE and CPMIDP02 CE are the insertion/deletion of83 and 5 base pairs(bp),and belong to the intergenic region of trn S-trn G and ndh J-ndh K.The amplified products of CPMIDP01 CE and CPMIDP02 CE primers contain 83 bp insertion sequence(fragment length is 339bp),do not contain 5bp insertion sequence(fragment length is 239bp),the analysis samples are S-type CMS materials.The detection methods were established using the twospecific primers based on different electrophoresis platforms,such as denaturing fluorescence capillary,non-denatured gel capillary,agarose electrophoresis platform.Moreover,the two primers have the potential to be compatible with more efficient KASP,Taq Man and other technical systems.In this research,in order to identify maize S-type CMS materials,a simple-fast,stable-efficient,low cost and innovative method was constructed by chloroplast InDel markers,and it will provide key technical support for the production of maize hybrids in China.