Isolation and characterization of kinetoplast DNA from bloodstream form of Trypanosoma brucei.
J Cell Biol
; 76(2): 293-309, 1978 Feb.
Article
em En
| MEDLINE
| ID: mdl-10605439
ABSTRACT
We have used restriction endonucleases PstI, EcoRI, HapII, HhaI, and S1 nuclease to demonstrate the presence of a large complex component, the maxi-circle, in addition to the major mini-circle component in kinetoplast DNA (kDNA) networks of Trypanosoma brucei (East African Trypanosomiasis Research Organization [EATRO] 427). Endonuclease PstI and S1 nuclease cut the maxi-circle at a single site, allowing its isolation in a linear form with a mol wt of 12.2 x 10(6), determined by electron microscopy. The other enzymes give multiple maxi-circle fragments, whose added mol wt is 12-13 x 10(6), determined by gel electrophoresis. The maxi-circle in another T. brucei isolate (EATRO 1125) yields similar fragments but appears to contain a deletion of about 0.7 x 10(6) daltons. Electron microscopy of kDNA shows the presence of DNA considerably longer than the mini-circle contour length (0.3 micron) either in the network or as loops extending from the edge. This long DNA never exceeds the maxi-circle length (6.3 microns) and is completely removed by digestion with endonuclease PstI. 5-10% of the networks are doublets with up to 40 loops of DNA clustered between the two halves of the mini-circle network and probably represent a division stage of the kDNA. Digestion with PstI selectively removes these loops without markedly altering the mini-circle network. We conclude that the long DNA in both single and double networks represents maxi-circles and that long tandemly repeated oligomers of mini-circles are (virtually) absent. kDNA from Trypanosoma equiperdum, a trypanosome species incapable of synthesizing a fully functional mitochondrion, contains single and double networks of dimensions similar to those from T. brucei but without any DNA longer than mini-circle contour length. We conclude that the maxi-circle of trypanosomes is the genetic equivalent of the mitochondrial DNA (mtDNA) of other organisms.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Trypanosoma brucei brucei
/
DNA de Cinetoplasto
Limite:
Animals
Idioma:
En
Revista:
J Cell Biol
Ano de publicação:
1978
Tipo de documento:
Article
País de afiliação:
Holanda