[Construction and characteristics of a cosmid library of genes of the bacterium Cornyebacterium glutamicum ATSS13032].
A representative genomic library of the Corynebacterium glutamicum ATCC 13032 genes in a cosmid vector Lorist6 was created. The cosmids contain inserts of bacterial DNA obtained by partial digestion with the Sau3A I restrictase. Five hundred and thirty individual primary recombinant clones were transferred into the wells of microtiter plates, where they are now being preserved. The average size of the bacterial DNA inserts determined via a sum of restriction fragment sizes of recombinant molecules is about 38 kb. The capacity of the obtained gene library is 8.4 equivalents of the C. glutamicum genome, i.e., every fragment of the genome is on average represented by eight clones and is presented in at least one clone with the probability > 99%. Clone grids (sets of recombinant clones located on the hybridization membrane in regular and reproducible order) were created. Specificity of the created clone library and its representativeness were confirmed experimentally by hybridization of clone grids with DNA probes corresponding to unique regions of the Corynebacterium genome. A plasmid containing the pheA prephenate dehydratase gene, olygonucleotide corresponding to the lysC gene, and the 21 RNA probe obtained from the insert ends in different cosmids were used as probes. The created set of clones allows the construction of a cosmid contig overlapping the C. glutamicum genome and a physical genetic map on its base.
Bukanov
N O
NO
Vavilov Institute of General Genetics, Russian Academy of Sciences, Moscow, Russia.
Nashchokina
O O
OO
Borinskaia
S A
SA
Lobashev
A V
AV
Fonshteĭn
M Iu
MIu
Gusiatiner
M M
MM
Debabov
V G
VG
Iankovskiĭ
N K
NK
D003352:Corynebacterium / Q000235:genetics*; D003360:Cosmids / Q000235:genetics*; D015723:Gene Library; D005798:Genes, Bacterial; D005822:Genetic Vectors; D016680:Genome, Bacterial; D015183:Restriction Mapping
Konstruirovanie i kharakteristika kosmidnoĭ biblioteki genov bakterii Corynebacterium glutamicum ATCC13032.