OBJECTIVE To understand the ctxB and rstR variations of toxigenic Vibrio cholerae (V.cholerae) O1 El Tor strains isolated from different provinces in China from 1961 to 2010. METHODS All 385 toxigenic V.cholerae O1 El Tor strains were selected, which were isolated in China between year 1961 and 2010. ctxB gene was amplified by PCR method and sequenced for further analysis. rstR was detected with PCR by using the genotype specific primers. RESULTS ctxB sequence analysis revealed that 52.5% (202/385) isolates carried ctxB(ET) and 47.5% (183/385) carried ctxB(class), namely Y(39) to H and I(68) to T substitutions which were specific to the classical biotype CT-B sequence. From 1961 to 1992, strains carrying ctxB(ET) were predominant and the proportion was as high as 98.4% (182/185). After 1993, strains carrying ctxB(class) were sharply increased. Especially during year 1993 to year 2005, 97.2% (174/179) of the isolated strains carried ctxB(class). Since 2006, resurgence of dominant strains carrying ctxB(ET) or co-existing of strains with ctxB(ET) or ctxB(class) was noticed. rstR genotype detection showed that 62.9% (242/385)of the tested strains carried the rstR(ET), while 6.8% (26/385) with rstR(class), and the remainings contained at least two types of rstR in different combination forms, among which rstR(ET)+rstR(class) combination were the most, accounting for 75.7% (75/99) . Similar to the ctxB, the distribution of rstR genotypes showed time specificity. From 1961 to 1992, strains carrying rstR(ET) predominated (87.0%, 161/185). After 1993, the diversity of rstR genotypes was observed accompanying by a sharp increase of strains containing other rstR genotype, such as rstR(class), rstR(env) and different combinations. There were separately 96% (25/26), 84% (63/75) and 18/18 strains containing rstR(class), rstR(ET)+rstR(class) and rstR(ET)+rstR(class)+rstR(env) isolated after 1993. CONCLUSION The distribution of different genotypes of ctxB and rstR showed obvious time-specificity, and there were various combining forms of rstR, reflecting the diversity of the genetic and evolutionary characteristics of Chinese V.cholerae isolates.