Next Article in Journal
Increased Susceptibility of Radiation-Induced Intestinal Apoptosis in SMP30 KO Mice
Previous Article in Journal
Ubiquitin-Conjugating Enzyme 9 Promotes Epithelial Ovarian Cancer Cell Proliferation in Vitro
Int. J. Mol. Sci. 2013, 14(6), 11072-11083; doi:10.3390/ijms140611072
Article

Construction of a Full-Length Enriched cDNA Library and Preliminary Analysis of Expressed Sequence Tags from Bengal Tiger Panthera tigris tigris

1,2,†
,
3,†
,
2
,
1
,
1
,
4
,
4
,
1
 and
1,*
1 Institute of Animal Science, Chinese Academy of Agricultural Sciences, Beijing 100193, China 2 Department of Bioscience, Bengbu Medical College, Bengbu 233000, China 3 The Northeast Tiger Wooden Land of Heilongjiang, Harbin 150028, China 4 College of Wildlife Resource, Northeast Forestry University, Harbin 150028, China These authors contributed equally to this work.
* Author to whom correspondence should be addressed.
Received: 21 November 2012 / Revised: 24 April 2013 / Accepted: 25 April 2013 / Published: 24 May 2013
(This article belongs to the Section Biochemistry, Molecular Biology and Biophysics)
View Full-Text   |   Download PDF [3105 KB, uploaded 19 June 2014]   |  

Abstract

In this study, a full-length enriched cDNA library was successfully constructed from Bengal tiger, Panthera tigris tigris, the most well-known wild Animal. Total RNA was extracted from cultured Bengal tiger fibroblasts in vitro. The titers of primary and amplified libraries were 1.28 × 106 pfu/mL and 1.56 × 109 pfu/mL respectively. The percentage of recombinants from unamplified library was 90.2% and average length of exogenous inserts was 0.98 kb. A total of 212 individual ESTs with sizes ranging from 356 to 1108 bps were then analyzed. The BLASTX score revealed that 48.1% of the sequences were classified as a strong match, 45.3% as nominal and 6.6% as a weak match. Among the ESTs with known putative function, 26.4% ESTs were found to be related to all kinds of metabolisms, 19.3% ESTs to information storage and processing, 11.3% ESTs to posttranslational modification, protein turnover, chaperones, 11.3% ESTs to transport, 9.9% ESTs to signal transducer/cell communication, 9.0% ESTs to structure protein, 3.8% ESTs to cell cycle, and only 6.6% ESTs classified as novel genes. By EST sequencing, a full-length gene coding ferritin was identified and characterized. The recombinant plasmid pET32a-TAT-Ferritin was constructed, coded for the TAT-Ferritin fusion protein with two 6× His-tags in N and C-terminal. After BCA assay, the concentration of soluble Trx-TAT-Ferritin recombinant protein was 2.32 ± 0.12 mg/mL. These results demonstrated that the reliability and representativeness of the cDNA library attained to the requirements of a standard cDNA library. This library provided a useful platform for the functional genome and transcriptome research of Bengal tigers.
Keywords: Bengal tiger; Panthera tigris tigris; fibroblast cell line; SMART cDNA library; expressed sequence tags Bengal tiger; Panthera tigris tigris; fibroblast cell line; SMART cDNA library; expressed sequence tags
This is an open access article distributed under the Creative Commons Attribution License (CC BY 3.0).
SciFeed

Share & Cite This Article

Further Mendeley | CiteULike
Export to BibTeX |
EndNote |
RIS
MDPI and ACS Style

Liu, C.; Liu, D.; Guo, Y.; Lu, T.; Li, X.; Zhang, M.; Ma, J.; Ma, Y.; Guan, W. Construction of a Full-Length Enriched cDNA Library and Preliminary Analysis of Expressed Sequence Tags from Bengal Tiger Panthera tigris tigris. Int. J. Mol. Sci. 2013, 14, 11072-11083.

View more citation formats

Related Articles

Article Metrics

For more information on the journal, click here

Comments

[Return to top]
Int. J. Mol. Sci. EISSN 1422-0067 Published by MDPI AG, Basel, Switzerland RSS E-Mail Table of Contents Alert