Molecular cloning, characterization and expression of Mn-superoxide dismutase from the rubber tree (Hevea brasiliensis)

Zhonghe Miao, John Gaynor

Research output: Contribution to journalArticleResearchpeer-review

48 Citations (Scopus)

Abstract

A genomic clone encoding manganese-containing superoxide dismutase (SOD; EC 1.15.1.1) was isolated from a Hevea brasiliensis genomic library made in λ phage EMBL3 by using a heterologous cDNA probe of MnSOD from Nicotiana plumbaginifolia. The nucleotide sequence of 4968 bp from the genomic clone was determined. Based on the putative translation initiation codon and stop codon, PCR primers were designed and utilized for cloning the full-length cDNA from total mRNA. Of the two distinct cDNAs of MnSOD isolated, MnSOD-A has a perfect match with exons of the nuclear gene, while MnSOD-B has a 90.2% homology and is 6 nucleotides longer than MnSOD-A in the putative transit peptide region. The nuclear gene comprises 6 exons and 5 introns, giving a total length of 3211 bp. The sequences of 1400 bp upstream of the initiation codon and 320 bp downstream of the stop codon were also determined. Southern analysis of genomic DNA from Hevea probed with a genomic fragment indicated there are at least two genes of MnSOD in Hevea. Northern blot analysis showed that MnSOD transcripts were present in all tissues examined (leaf, petiole, root, latex, callus) with young leaves showing the highest levels in intact plants. The transcript level in embryogenic callus was nearly 50-fold higher than in mature leaves. In addition, transcripts of MnSOD could be induced 3- to 5-fold in response to sucrose, ethephon and Murashige-Skoog salts.

Original languageEnglish
Pages (from-to)267-277
Number of pages11
JournalPlant Molecular Biology
Volume23
Issue number2
DOIs
StatePublished - 1 Oct 1993

Fingerprint

Hevea
Hevea brasiliensis
Molecular Cloning
Superoxide Dismutase
molecular cloning
superoxide dismutase
Initiator Codon
Complementary DNA
Terminator Codon
Bony Callus
genomics
start codon
stop codon
Exons
Clone Cells
Genes
exons
callus
Genomic Library
Latex

Keywords

  • cDNA clone
  • genomic clone
  • MnSOD
  • nucleotide sequence
  • PCR

Cite this

@article{dd97e429649a4a37bfd1a58541f715f6,
title = "Molecular cloning, characterization and expression of Mn-superoxide dismutase from the rubber tree (Hevea brasiliensis)",
abstract = "A genomic clone encoding manganese-containing superoxide dismutase (SOD; EC 1.15.1.1) was isolated from a Hevea brasiliensis genomic library made in λ phage EMBL3 by using a heterologous cDNA probe of MnSOD from Nicotiana plumbaginifolia. The nucleotide sequence of 4968 bp from the genomic clone was determined. Based on the putative translation initiation codon and stop codon, PCR primers were designed and utilized for cloning the full-length cDNA from total mRNA. Of the two distinct cDNAs of MnSOD isolated, MnSOD-A has a perfect match with exons of the nuclear gene, while MnSOD-B has a 90.2{\%} homology and is 6 nucleotides longer than MnSOD-A in the putative transit peptide region. The nuclear gene comprises 6 exons and 5 introns, giving a total length of 3211 bp. The sequences of 1400 bp upstream of the initiation codon and 320 bp downstream of the stop codon were also determined. Southern analysis of genomic DNA from Hevea probed with a genomic fragment indicated there are at least two genes of MnSOD in Hevea. Northern blot analysis showed that MnSOD transcripts were present in all tissues examined (leaf, petiole, root, latex, callus) with young leaves showing the highest levels in intact plants. The transcript level in embryogenic callus was nearly 50-fold higher than in mature leaves. In addition, transcripts of MnSOD could be induced 3- to 5-fold in response to sucrose, ethephon and Murashige-Skoog salts.",
keywords = "cDNA clone, genomic clone, MnSOD, nucleotide sequence, PCR",
author = "Zhonghe Miao and John Gaynor",
year = "1993",
month = "10",
day = "1",
doi = "10.1007/BF00029003",
language = "English",
volume = "23",
pages = "267--277",
journal = "Plant Molecular Biology",
issn = "0167-4412",
publisher = "Springer Netherlands",
number = "2",

}

Molecular cloning, characterization and expression of Mn-superoxide dismutase from the rubber tree (Hevea brasiliensis). / Miao, Zhonghe; Gaynor, John.

In: Plant Molecular Biology, Vol. 23, No. 2, 01.10.1993, p. 267-277.

Research output: Contribution to journalArticleResearchpeer-review

TY - JOUR

T1 - Molecular cloning, characterization and expression of Mn-superoxide dismutase from the rubber tree (Hevea brasiliensis)

AU - Miao, Zhonghe

AU - Gaynor, John

PY - 1993/10/1

Y1 - 1993/10/1

N2 - A genomic clone encoding manganese-containing superoxide dismutase (SOD; EC 1.15.1.1) was isolated from a Hevea brasiliensis genomic library made in λ phage EMBL3 by using a heterologous cDNA probe of MnSOD from Nicotiana plumbaginifolia. The nucleotide sequence of 4968 bp from the genomic clone was determined. Based on the putative translation initiation codon and stop codon, PCR primers were designed and utilized for cloning the full-length cDNA from total mRNA. Of the two distinct cDNAs of MnSOD isolated, MnSOD-A has a perfect match with exons of the nuclear gene, while MnSOD-B has a 90.2% homology and is 6 nucleotides longer than MnSOD-A in the putative transit peptide region. The nuclear gene comprises 6 exons and 5 introns, giving a total length of 3211 bp. The sequences of 1400 bp upstream of the initiation codon and 320 bp downstream of the stop codon were also determined. Southern analysis of genomic DNA from Hevea probed with a genomic fragment indicated there are at least two genes of MnSOD in Hevea. Northern blot analysis showed that MnSOD transcripts were present in all tissues examined (leaf, petiole, root, latex, callus) with young leaves showing the highest levels in intact plants. The transcript level in embryogenic callus was nearly 50-fold higher than in mature leaves. In addition, transcripts of MnSOD could be induced 3- to 5-fold in response to sucrose, ethephon and Murashige-Skoog salts.

AB - A genomic clone encoding manganese-containing superoxide dismutase (SOD; EC 1.15.1.1) was isolated from a Hevea brasiliensis genomic library made in λ phage EMBL3 by using a heterologous cDNA probe of MnSOD from Nicotiana plumbaginifolia. The nucleotide sequence of 4968 bp from the genomic clone was determined. Based on the putative translation initiation codon and stop codon, PCR primers were designed and utilized for cloning the full-length cDNA from total mRNA. Of the two distinct cDNAs of MnSOD isolated, MnSOD-A has a perfect match with exons of the nuclear gene, while MnSOD-B has a 90.2% homology and is 6 nucleotides longer than MnSOD-A in the putative transit peptide region. The nuclear gene comprises 6 exons and 5 introns, giving a total length of 3211 bp. The sequences of 1400 bp upstream of the initiation codon and 320 bp downstream of the stop codon were also determined. Southern analysis of genomic DNA from Hevea probed with a genomic fragment indicated there are at least two genes of MnSOD in Hevea. Northern blot analysis showed that MnSOD transcripts were present in all tissues examined (leaf, petiole, root, latex, callus) with young leaves showing the highest levels in intact plants. The transcript level in embryogenic callus was nearly 50-fold higher than in mature leaves. In addition, transcripts of MnSOD could be induced 3- to 5-fold in response to sucrose, ethephon and Murashige-Skoog salts.

KW - cDNA clone

KW - genomic clone

KW - MnSOD

KW - nucleotide sequence

KW - PCR

UR - http://www.scopus.com/inward/record.url?scp=0027672185&partnerID=8YFLogxK

U2 - 10.1007/BF00029003

DO - 10.1007/BF00029003

M3 - Article

VL - 23

SP - 267

EP - 277

JO - Plant Molecular Biology

JF - Plant Molecular Biology

SN - 0167-4412

IS - 2

ER -