Rolling circle amplification of genomic templates for inverse PCR (RCA-GIP): a method for 5'- and 3'-genome walking without anchoring

被引:24
作者
Tsaftaris, Athanasios [1 ,2 ]
Pasentzis, Konstantinos [1 ]
Argiriou, Anagnostis [1 ]
机构
[1] Ctr Res & Technol Hellas, Inst Agrobiotechnol, Thessaloniki 57001, Thermi, Greece
[2] Aristotle Univ Thessaloniki, GR-54006 Thessaloniki, Greece
关键词
Genome walking; Inverse PCR; phi; 29; Rolling cycle amplification; MADS-BOX GENES; CROCUS-SATIVUS L; CLONING; ARABIDOPSIS; EXPRESSION;
D O I
10.1007/s10529-009-0128-9
中图分类号
Q81 [生物工程学(生物技术)]; Q93 [微生物学];
学科分类号
071005 ; 0836 ; 090102 ; 100705 ;
摘要
We have devised an improved method of genome walking, named rolling circle amplification of genomic templates for Inverse PCR (RCA-GIP). The method is based on the generation of circular genomic DNA fragments, followed by rolling circle amplification of the circular genomic DNA using I center dot 29 DNA polymerase without need for attachment of anchor sequences. In this way from the circular genomic DNA fragments, after RCA amplification, a large amount of linear concatemers is generated suitable for Inverse PCR template that can be amplified, sequenced or cloned allowing the isolation of the 3'- and 5'- of unknown ends of genomic sequences. To prove the concept of the proposed methodology, we used this procedure to isolate the promoter regions from different species. Herein as an example we present the isolation of four promoter regions from Crocus sativus, a crop cultivated for saffron production.
引用
收藏
页码:157 / 161
页数:5
相关论文
共 14 条
[1]   trans meets cis in MADS science [J].
de Folter, Stefan ;
Angenent, Gerco C. .
TRENDS IN PLANT SCIENCE, 2006, 11 (05) :224-231
[2]   Heterotopic expression of B-class floral homeotic genes PISTILLATA/GLOBOSA supports a modified model for crocus (Crocus sativus L.) flower formation [J].
Kalivas, Apostolos ;
Pasentsis, Konstantinos ;
Polidoros, Alexios N. ;
Tsaftaris, Athanasios S. .
DNA SEQUENCE, 2007, 18 (02) :120-130
[3]   Rolling circle amplification-RACE:: a method for simultaneous isolation of 5′ and 3′ cDNA ends from amplified cDNA templates [J].
Polidoros, Alexios N. ;
Pasentsis, Konstantinos ;
Tsaftaris, Athanasios S. .
BIOTECHNIQUES, 2006, 41 (01) :35-+
[4]  
SATO M, 2004, DNA NUCL ACIDS S SER, P147
[5]   LONG-RANGE WALKING TECHNIQUES IN POSITIONAL CLONING STRATEGIES [J].
STUBBS, L .
MAMMALIAN GENOME, 1992, 3 (03) :127-142
[6]   Binding site selection for the plant MADS domain protein AGL15 -: An in vitro and in vivo study [J].
Tang, WN ;
Perry, SE .
JOURNAL OF BIOLOGICAL CHEMISTRY, 2003, 278 (30) :28154-28159
[7]   CHROMOSOME LANDING - A PARADIGM FOR MAP-BASED GENE CLONING IN PLANTS WITH LARGE GENOMES [J].
TANKSLEY, SD ;
GANAL, MW ;
MARTIN, GB .
TRENDS IN GENETICS, 1995, 11 (02) :63-68
[8]   Tepal formation and expression pattern of B-class paleoAP3-like MADS-box genes in crocus (Crocus sativus L.) [J].
Tsaftaris, AS ;
Polidoros, AN ;
Pasentsis, K ;
Kalivas, A .
PLANT SCIENCE, 2006, 170 (02) :238-246
[9]   Isolation of a differentially spliced C-type flower specific AG-like MADS-box gene from Crocus sativus and characterization of its expression [J].
Tsaftaris, AS ;
Pasentsis, K ;
Polidoros, AN .
BIOLOGIA PLANTARUM, 2005, 49 (04) :499-504
[10]   Isolation of three homologous AP1-like MADS-box genes in crocus (Crocus sativus L.) and characterization of their expression [J].
Tsaftaris, AS ;
Pasentsis, K ;
Iliopoulos, I ;
Polidoros, AN .
PLANT SCIENCE, 2004, 166 (05) :1235-1243