1022-7954/01/3705- $25.00 © 2001
Russian Journal of Genetics, Vol. 37, No. 5, 2001, pp. 494–499. Translated from Genetika, Vol. 37, No. 5, 2001, pp. 610–616.
Original Russian Text Copyright © 2001 by Andronov, Roumiantseva, Simarov.
Many researchers have considered plasmids and IS
elements as “additional” elements that need not be
present in the genome [1, 2]. Nevertheless, plasmids
often compose the largest portion of the genome in
nodule bacteria. Thus, plasmid DNA may constitute up
to 50% of the rhizobial genome , and the number of
cryptic (nonsymbiotic) plasmids can reach 10 . At
present, genetic determinants located in cryptic plas-
mids have been shown to affect opportunities of strains
for adapting to the environment and the interaction with
the host plant .
The existence of many repeated sequences, includ-
ing IS elements, is characteristic of the
genome [6–8]. Bacterial IS elements do not contain
genetic determinants, except for those that are required
for their own transposition . Some researchers
believe that it is the ability of IS elements to induce a
variety of genomic rearrangements that ensures a high
level of genetic diversity in
[6, 10]. To date,
more than ten various IS elements have been detected
in nodule bacteria of alfalfa, and the IS
ment is studied in most detail. This IS element is unique
in that it was detected in the majority of natural strains
of different geographical ori-
gin and has a high copy number [7, 11, 12].
The purpose of this work is to examine a variety of
plasmid proﬁles and IS
2011-2 ﬁngerprints of natu-
strains isolated in two geographically dis-
tant sites from the soil under different species of wild
plants, alfalfa and legume. Additionally, it has been of
interest to compare data derived from the analysis of
plasmid proﬁles in strains and of IS ﬁngerprints with
the results of RFLP analysis of the same strains con-
ducted previously .
MATERIALS AND METHODS
Strains used in this work.
We isolated 56 natural
(Table 1) from soil samples in two
ﬁeld sites (A and B) located in the Irkutsk oblast at a
distance of 80 km from each other. In each ﬁeld site,
samples were collected from the soil under two differ-
ent wild host plants, alfalfa (
) . We used two
strains with the known genetic characteris-
tics: Rm2011  and MVII .
Analysis of plasmid proﬁle in S. meliloti strains.
Plasmids were identiﬁed by the Eckhardt method 
modiﬁed for rhizobia .
Total bacterial DNA
was isolated according to the standard method .
Total DNA was hydrolyzed with the
enzyme. Restriction DNA fragments were separated by
electrophoresis and transferred onto a nylon membrane
by the standard technique . A PCR-ampliﬁed
2011-2 element  labeled with digoxigenin
was used for Southern blotting. Nonradioactive hybrid-
ization and detection were conducted in accordance
with recommendations of the manufacturer (Boeringer
Statistical treatment of data.
For statistical analysis,
test for similarity between variation ranges was
used . Analysis of correlation (linkage) between the
plasmid type and chromosomal RFLP type was con-
ducted by means of the Arlequine 1.1 program .
Genetic Diversity of a Natural Population
Revealed in Analysis
of Cryptic Plasmids and IS
E. E. Andronov, M. L. Roumiantseva, and B. V. Simarov
All-Russia Research Institute of Agricultural Microbiology, Russian Academy of Agricultural Sciences,
St. Petersburg, Pushkin, 196608 Russia; fax: (812) 470-43-62; e-mail: email@example.com
Received October 30, 2000
—Fifty-six natural strains of alfalfa nodule bacteria were isolated from samples of the soil under wild
legume and alfalfa in two different ﬁeld sites of Irkutsk oblast. Based on the results of analysis of plasmid pro-
ﬁle, 11 different types of strains were detected, and 43 types were identiﬁed based on the results of hybridiza-
tion with the insertion sequence element IS
2011-2. Signiﬁcant differences were found in the plasmid proﬁle
and IS ﬁngerprints between strains isolated from the soil under alfalfa and the soil under legume. In contrast,
strains growing at some distance from each other differed only in the IS ﬁngerprints. From a comparison of
results obtained in the assessment of plasmid proﬁle and in analysis of IS ﬁngerprints with results of RFLP anal-
ysis in strains, the conclusion about the transference of cryptic plasmids between strains and genetic rearrange-
ments in strains of this population was drawn.