Characterization of Genomic Inheritance of Intergeneric Hybrids between Ascocenda and Phalaenopsis Cultivars by GISH, PCR-RFLP and RFLP.

Wen-Lin Liu,Chi-Chu Tsai,Ya-Zhu Ko,Chang-Hung Chou,I-Szu Weng,Yu-Chung Chiang,Huei-Chuan Shih,Tzen-Yuh Chiang

doi:10.1371/journal.pone.0153512

Abstract

BackgroundThe intergeneric hybrids between Ascocenda John De Biase ‘Blue’ and Phalaenopsis Chih Shang's Stripes have been generated to introduce the blue color into the Phalaenopsis germplasm in prior study. In order to confirm the inheritance in hybrid progenies, genomic in situ hybridization (GISH) and restriction fragment length polymorphism (RFLP) analysis were conducted to confirm the intergeneric hybridization status.Methods/ResultsGISH analysis showed the presence of both maternal and paternal chromosomes in the cells of the putative hybrids indicating that the putative hybrid seedlings were intergeneric hybrids of the two parents. Furthermore, twenty-seven putative hybrids were randomly selected for DNA analysis, and the external transcribed spacer (ETS) regions of nrDNA were analyzed using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and RFLP analyses to identify the putative hybrids. RFLP analysis showed that the examined seedlings were intergeneric hybrids of the two parents. However, PCR-RFLP analysis showed bias to maternal genotype.ConclusionsBoth GISH and RFLP analyses are effective detection technology to identify the intergeneric hybridization status of putative hybrids. Furthermore, the use of PCR-RFLP analysis to identify the inheritance of putative hybrids should be carefully evaluated.

Highlights

Comprising approximately 35,000 species, Orchidaceae is considered the largest family of flowering plants in the world [1]
The use of PCR-restriction fragment length polymorphism (RFLP) analysis to identify the inheritance of putative hybrids should be carefully evaluated
Vandaceous orchids include Vanda, Ascocenda, Phalaenopsis, Renanthera, Rhynchostylis and Aerides, which are characterized by a monopodial growth habit and are mainly found in tropical Asia [2]

Summary

Introduction

Comprising approximately 35,000 species, Orchidaceae is considered the largest family of flowering plants in the world [1]. Vandaceous orchids include Vanda, Ascocenda, Phalaenopsis, Renanthera, Rhynchostylis and Aerides, which are characterized by a monopodial growth habit and are mainly found in tropical Asia [2]. Ascocenda John De Biase ‘Blue’ is an intergeneric hybrid between Ascocentrum and Vanda species [6]. P. Chih Shang's Stripes is an interspecific hybrid, and its genetic background includes species of P. amboinensis, P. lueddemanniana, P. amabilis, P. aphrodite, P. equestris, P. schilleriana, P. sanderiana, and P. stuartiana [6]. Chih Shang's Stripes have been generated to introduce the blue color into the Phalaenopsis germplasm in prior study [11]. The intergeneric hybrids between Ascocenda John De Biase ‘Blue’ and Phalaenopsis Chih Shang's Stripes have been generated to introduce the blue color into the Phalaenopsis germplasm in prior study. In order to confirm the inheritance in hybrid progenies, genomic in situ hybridization (GISH) and restriction fragment length polymorphism (RFLP) analysis were conducted to confirm the intergeneric hybridization status

Methods

Results

Conclusion