The Cloning and Fluorescence In situ Hybridization Analysis of Cotton Telomere Sequence

doi:10.1016/S1671-2927(00)8672

Journal of Integrative Agriculture ›› 2012, Vol. 12 ›› Issue (9): 1417-1423.DOI: 10.1016/S1671-2927(00)8672

The Cloning and Fluorescence In situ Hybridization Analysis of Cotton Telomere Sequence

LING Jian, CHENG Hua, LIU Fang, SONG Guo-li, WANG Chun-ying, LI Shao-hui, ZHANG Xiang-di, WANG Yu-hong, WANG Kun-bo

1.State Key Laboratory of Cotton Biology/Cotton Research Institute, Chinese Academy of Agricultural Sciences, Anyang 455000, P.R.China
2.Anyang Institute of Technology, Anyang 455000, P.R.China

收稿日期:2011-05-09 出版日期:2012-09-01 发布日期:2012-09-09
通讯作者: Correspondence WANG Kun-bo, Tel: +86-372-2562203, Fax: +86-372-2525377, E-mail: wkbcri@cricaas.com.cn
基金资助:
This work was supported by the National Natural Science Foundation of China (30170501).

The Cloning and Fluorescence In situ Hybridization Analysis of Cotton Telomere Sequence

LING Jian, CHENG Hua, LIU Fang, SONG Guo-li, WANG Chun-ying, LI Shao-hui, ZHANG Xiang-di, WANG Yu-hong, WANG Kun-bo

1.State Key Laboratory of Cotton Biology/Cotton Research Institute, Chinese Academy of Agricultural Sciences, Anyang 455000, P.R.China
2.Anyang Institute of Technology, Anyang 455000, P.R.China

Received:2011-05-09 Online:2012-09-01 Published:2012-09-09
Contact: Correspondence WANG Kun-bo, Tel: +86-372-2562203, Fax: +86-372-2525377, E-mail: wkbcri@cricaas.com.cn
Supported by:
This work was supported by the National Natural Science Foundation of China (30170501).

摘要/Abstract

摘要： Telomeres form the ends of eukaryotic chromosomes and serve as protective caps that keep chromosomes structure independency and completeness. The first plant telomere DNA was isolated from Arabidopsis thaliana and was shown to have tandemly repeated sequence 5´-TTTAGGG-3´. The Arabidopsis-type telomere has been found in many plants, but several reports indicate that this sequence is absent in some plants. Up to now, no research has been conducted on the telomere of cotton. In this paper, the Arabidopsis-type telomere sequence was amplified and cloned using the primers designed based on the fragment containing telomere sequence in an Arabidopsis bacterial artificial chromosome (BAC). Fluorescence in situ hybridization (FISH) with cotton metaphase chromosomes using the Arabidopsis-type telomere sequence as probes indicated that the signals were located at all chromosome ends of seven diploid and two tetraploid cotton species with different signal intensities among chromosome complements of different cotton species, even between long and short arms of the same chromosome. To identify the signals of FISH, the genome DNA of Xinhai 7, a cultivar of Gossypium barbadense, digested by BAL-31 nuclease was introduced in this study. The result of BAL-31 digestion indicated that the hybridization signals of FISH represent the outermost DNA sequence of each cotton chromosomes. So we first proved that the telomeric repeats of cotton cross-hybridize with that of Arabidopsis. The results of terminal restriction fragment (TRF) showed significant variation in telomere length among cotton species. The telomere length of cultivated cotton was close to 20 kb and was larger than those of wild cotton species whose telomere length ranged from 6 to 20 kb.

关键词: cotton, fluorescent in situ hybridization (FISH), telomere, terminal restriction fragment (TRF)

Abstract: Telomeres form the ends of eukaryotic chromosomes and serve as protective caps that keep chromosomes structure independency and completeness. The first plant telomere DNA was isolated from Arabidopsis thaliana and was shown to have tandemly repeated sequence 5´-TTTAGGG-3´. The Arabidopsis-type telomere has been found in many plants, but several reports indicate that this sequence is absent in some plants. Up to now, no research has been conducted on the telomere of cotton. In this paper, the Arabidopsis-type telomere sequence was amplified and cloned using the primers designed based on the fragment containing telomere sequence in an Arabidopsis bacterial artificial chromosome (BAC). Fluorescence in situ hybridization (FISH) with cotton metaphase chromosomes using the Arabidopsis-type telomere sequence as probes indicated that the signals were located at all chromosome ends of seven diploid and two tetraploid cotton species with different signal intensities among chromosome complements of different cotton species, even between long and short arms of the same chromosome. To identify the signals of FISH, the genome DNA of Xinhai 7, a cultivar of Gossypium barbadense, digested by BAL-31 nuclease was introduced in this study. The result of BAL-31 digestion indicated that the hybridization signals of FISH represent the outermost DNA sequence of each cotton chromosomes. So we first proved that the telomeric repeats of cotton cross-hybridize with that of Arabidopsis. The results of terminal restriction fragment (TRF) showed significant variation in telomere length among cotton species. The telomere length of cultivated cotton was close to 20 kb and was larger than those of wild cotton species whose telomere length ranged from 6 to 20 kb.

Key words: cotton, fluorescent in situ hybridization (FISH), telomere, terminal restriction fragment (TRF)

LING Jian, CHENG Hua, LIU Fang, SONG Guo-li, WANG Chun-ying, LI Shao-hui, ZHANG Xiang-di, WANG Yu-hong, WANG Kun-bo. The Cloning and Fluorescence In situ Hybridization Analysis of Cotton Telomere Sequence[J]. Journal of Integrative Agriculture, 2012, 12(9): 1417-1423.

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The Cloning and Fluorescence In situ Hybridization Analysis of Cotton Telomere Sequence

The Cloning and Fluorescence In situ Hybridization Analysis of Cotton Telomere Sequence

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