Molecular Biology and Genetics
 

Plant Biotechnology

Electronic Journal of Biotechnology ISSN: 0717-3458 Vol. 11 No. 1, Issue of January 15, 2008
© 2008 by Pontificia Universidad Católica de Valparaíso -- Chile Received May 29, 2007 / Accepted August 21, 2007
DOI: 10.2225/vol11-issue1-fulltext-11
RESEARCH ARTICLE

Identification of differentially expressed genes in developing cotton fibers(Gossypium hirsutum L) through differential display

Saima Iqbal
Plant Biotechnology Division
National Institute for Biotechnology and Genetic Engineering
Jhang Road, Faisalabad, Pakistan
Tel: 92 41 2651475 ext 263
Fax: 92 41 2651472
E-mail: saimaqbl@yahoo.com

Aftab Bashir*
Plant Biotechnology Division
National Institute for Biotechnology and Genetic Engineering
P.O. Box 577, Jhang road
Faisalabad, Pakistan
Tel: 92 41 2651475 79 ext 232
Fax: 92 41 2651472
E-mail: aftabbashir@nibge.org

Hafiza Masooma Naseer
Plant Biotechnology Division
National Institute for Biotechnology and Genetic Engineering
Jhang Road, Faisalabad, Pakistan
Tel: 92 41 2651475 ext 233
Fax: 92 41 2651472
E-mail: masoomasajid@yahoo.com

Moddassir Ahmed
Plant Biotechnology Division
National Institute for Biotechnology and Genetic Engineering
Jhang Road, Faisalabad, Pakistan
Tel: 92 41 2651475 ext 262
Fax: 92 41 2651472
E-mail: moddassir@nibge.org

Kauser A. Malik
Food and Agriculture
Planning Commission
P-Block, Room # 404
Pak Secretariat
Islamabad, Pakistan
Tel: 92 51 9201974
Fax: 92 41 9206993
E-mail: kamalik@comsats.net.pk

*Corresponding author

Financial support: This work was supported by the Pakistan Academy of Sciences, Higher Education Commission (Pakistan) PhD student grant and ASPL-II project of MinFAL (Ministry of Food Agriculture and Live Stock).

Keywords: cotton fiber, differentially expressed transcripts, Gossypium hirsutum, non-radioactive.

Abbreviations:

ARP: acidic ribosomal protein
CHS: chalcone synthase
DDRT-PCR: differential display reverse transcriptase polymerase chain reaction
DETs: differentially expressed transcripts
DPA: days post anthesis
ESTs: expressed sequence tags
MBD: methyl CpG binding domain
OPT: oligopeptide transporter protein
PCR: polymerase chain reaction
PG: polygalacturonase enzyme
RRF: ribosome recycling factor
SPL: squamosa promoter binding protein

Abstract   Full Text

Cotton fibers are differentiated, non-dividing cells that originate from the epidermal layer of developing ovules. To identify genes involved in cotton fiber development, we performed non-radioactive differential display reverse transcriptase PCR (DDRT-PCR) on the purified mRNA. This technique was tested on mRNA isolated from five different developmental stages of cotton fiber including 0, 5, 10, 15 and 20 DPA (days after pollination). The mRNA purified from total RNA was reversibly transcribed using three anchored oligo-dT primers. Polymerase chain reaction (PCR) amplification of each cDNA preparation was carried out in combination with seven arbitrary primers. The amplified products were resolved on 1% agarose gel containing ethidium bromide. DNA was extracted from seventeen differentially expressed bands and cloned in pTZ57R/T vector. The sequencing and BLAST search analysis indicated that 12 of the differentially expressed genes matched the previously characterized genes, while 3 of them matched the uncharacterized sequences of cotton fiber expressed sequence tags (ESTs) reported previously to be associated with cotton fiber and 2 of the clones had homology with putative proteins. The technique can be used to efficiently identify differentially expressed genes and can be expanded to large scale studies by increasing the number of random decamers.

Supported by UNESCO / MIRCEN network