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Zoological Research
Kunming Institute of Zoology, Chinese Academy of Sciences
ISSN: 2095-8137
Vol. 29, No. 3, 2008, pp. 245-252
Bioline Code: zr08038
Full paper language: Chinese
Document type: Research Article
Document available free of charge

Zoological Research, Vol. 29, No. 3, 2008, pp. 245-252

 en Construction Fractional Genomic Libraries and Screening Microsatellites DNA of Esox reieherti check for this species in other resources Dybowski
WANG, Hong-zhe; YIN, Qian-qian; FENG, Zhi-gang; LI, Da-yu; SUN, Xiao-wen & LI, Chan

Abstract

Esox reieherti check for this species in other resources Dybowsk genomic microsatellites were developed by using enrichment protocols combined with radioactive hybridization protocol. Four hundred to nine hundred base pair fragments were selected for the whole genome. DNA PCR amplification after digestion with restriction endonuclease Sau 3AI, and (CA)12, (GA)12 probes marked with biotin were used for microsatellite DNA enrichment. The product fragments were connected with carrier pGEM-T and transferred into DH5α Escherichia coli check for this species in other resources competent cells, and radioactive isotope probes marked with γ-32 P were used for the second hybridization. As a result, a total of 1600 bacteria were obtained in the microsatellite genomic libraries, positive clones accounted for 90.91% before hybridization and 81.25% after hybridization, amounting to 1300. One hundred and ninety-six positive clones were selected for sequencing, and 192 clones included microsatellite sequences. The microsatellite sequences obtained, mono-nucleotide, quad-nucleotide and quint-nucleotide repeat motifs were observed beside double-base-pairs CA/GT, GA/CT. Seventy primers were designed according to the flanking sequences by using software Primer Premier 5.0, and 32 primers were selected to be synthesized. After optimizing PCR reaction conditions, 28 primers were amplified and produced clear purpose bands. The aim of our research was to promote the development and utilization of E. reieherti genomic resource, and lay the foundation for optimizing E. reieherti breeding strain in order to detect the genetic diversity and construct a genetic map.

Keywords
Esox reieheri Dybowsk; Microsatellite DNA; Magnetic beads enrichment protocol

 
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Alternative site location: http://www.zoores.ac.cn/

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