Screening of different AFLP fragments between near-isogenic lines of male sterile and fertile Salvia miltiorrhiza and their comparison analysis

Abstract

There is distinctive advantage of using male sterile lines to breed new cultivar and produce hybrids, when compared with general breeding method on yield and quality. In our previous work, near-isogenic lines (NILs) of male sterile and fertile Salvia miltiorrhiza have been obtained through continuous hybridization in many years. In this investigation, 378 primer combination were screened by using AFLP and BSA technique, in which 26 markers amplified from seven primers were found to tightly link to male sterile gene. Based on these markers, two linkage genetic maps were constructed. A 2 027,2 028 bp fragment was amplifed from NILs of fertile and sterile S. miltiorrhiza, respectively, using genome walking technique and previous E11/M4-208 marker as template. Four base mutations were found in intron when comparing both fragments. Among all different markers between NILs of male sterile and fertile S. miltiorrhiza, four was found to have 100% identities to chromosome 1, 3 and 5 of Arabidopsis, namely, E01/M09-418, E05/M13-308, E05/M04-750 and E01/M01-204. The E01/M09-418 marker was very close to male sterile gene of S. miltiorrhiza with distance of 2.1 cM, which also had 100% identities to male sterile gene MS2 in Arabidopsis. Both were distributed in chromosome 3 of Arabidopsis. The 2 028 bp fragment also had 100% identities to MS2 gene. Another E05/M04-750 marker that had 100% identities to chromosome 5 of Arabidopsis was found to have high identities to POP085-M05 gene of poplars and low affinity calcium antiporter CAX2 of Arabidopsis with very low E-value. The constructed genetic map and differential fragments with potential functions found in this study provide a solid foundation to lock male sterile genes in S. miltiorrhiza genome and to discover their functions.