Gene cloning, expression and purification of human mitochondrial tRNALeu(UUR) and its mutant

Han Weiguo, Chen Li, Liu Jing, Zha Xiliang, Jin Youxin, Wang Debao

Research output: Contribution to journalArticle

Abstract

Genes of human mitochondrial tRNALeu(UUR) (mtRNALeu(UUR)) and its mutant (mtRNALeu(M)) were synthesized and inserted into the plasmid pGEM-9Zf(-) respectively. E.coli JM 109 was transformed by the recombinant plasmids containing the target genes. The mtRNALeu(UUR) and mtRNALeu(M) were expressed up to 19.10% and 17.76% of total small RNA respectively. They were purified to 54% homogeneity by DEAE-sepharose-CL4B column chromatography and finally repurified by 15% PAGE/urea. Their kinetic parameters for E.coli LeuRS were measured. The results showed that the value of kcal/ Km of mtRNALeu(M) was about one fifth of that of mtRNALeu(UUR) and indicated the leucine acceptability of mtRNALeu(M) was much lower than that of mtRNALeu(UUR).

Original languageEnglish (US)
JournalScience in China, Series C: Life Sciences
Volume44
Issue number2
StatePublished - Apr 2001
Externally publishedYes

Fingerprint

RNA, Transfer, Leu
Cloning
Purification
purification
Organism Cloning
molecular cloning
Genes
Gene Expression
mutants
gene
plasmid
plasmids
Escherichia coli
agarose
homogeneity
Plasmids
urea
leucine
chromatography
RNA

Keywords

  • Expression
  • Gene cloning
  • Human mitochondrial tRNA
  • Mutation
  • Purification

ASJC Scopus subject areas

  • Agricultural and Biological Sciences(all)
  • Agricultural and Biological Sciences (miscellaneous)

Cite this

Gene cloning, expression and purification of human mitochondrial tRNALeu(UUR) and its mutant. / Weiguo, Han; Li, Chen; Jing, Liu; Xiliang, Zha; Youxin, Jin; Debao, Wang.

In: Science in China, Series C: Life Sciences, Vol. 44, No. 2, 04.2001.

Research output: Contribution to journalArticle

Weiguo, Han ; Li, Chen ; Jing, Liu ; Xiliang, Zha ; Youxin, Jin ; Debao, Wang. / Gene cloning, expression and purification of human mitochondrial tRNALeu(UUR) and its mutant. In: Science in China, Series C: Life Sciences. 2001 ; Vol. 44, No. 2.
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AU - Youxin, Jin

AU - Debao, Wang

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