Screening of Sex in Asparagus at Early Growth Stages

Abstract

<jats:p>To determine the sex of asparagus (<jats:italic>Asparagus officinalis</jats:italic>) at the seedling stage, an easy, economical, and reliable method was developed. We used a modified single-step DNA extraction protocol, which resulted in a crude extract containing sufficient genomic DNA for use as a template. The male-specific marker (Asp1-T7sp) is a dominant marker and may lead to false negatives caused by an incomplete reaction; therefore, a multiplex polymerase chain reaction (PCR) was developed using a ribosomal RNA gene marker. The resulting banding pattern distinguished males from females without false negatives. To determine the best tissue for extraction of template DNA, phylloclades (a specialized stem that resembles and functions like a leaf) or root tips of individual asparagus plants were collected and weighed. A 4.0-mg phylloclade sample or a 0.8-mg root sample provided sufficient DNA for PCR analysis of asparagus. Root excision at day 19 did not affect subsequent growth of asparagus seedlings after 28 days. The method can determine the sex of asparagus at day 19 after seeding. A combination of single-step DNA extraction from root tips and multiplex PCR made for a simple and reliable screening method.</jats:p>

Effects of Cultivar and Cropping Type on the Growth and Yield of Female and Male Asparagus Plants

“…It should also be noted that the sex identification process for plants is time-consuming and can be costly because it involves DNA extraction and polymerase chain reaction analysis. Although the development of simpler sex identification methods is ongoing (Ii et al 2012), the cost of such methods must be considered when determining the cost-effectiveness of female plant production for each cropping type, cultivar, and region. The development of a simple and low-cost method for sex identification would be beneficial.…”

Establishment of male‐specific sequence‐tagged site markers in Asparagus officinalis: An efficient tool for sex identification

“…Our findings confirmed that Asp1‐T7sp marker was failed to amplify male‐specific fragments in several tested cultivars as compared with our newly developed markers. Several other studies also attempted to develop different molecular marker for sex identification in asparagus (Gebler et al, 2007; Ii, Uno, et al, 2012); however, the sensitivity and specificity of such DNA markers in asparagus still remained debatable.…”

“…In a recent study, a new co‐dominant STS marker for sex identification in asparagus was developed (Stone et al, 2017) but remained unsuccessful in distinguishing male and female plants in tetraploid asparagus. Similarly, a male‐specific STS marker MSSTS710 (Ii, Uno, et al, 2012) was demonstrated to distinguish male and female plants in garden asparagus; however, it is not successful in some related species of asparagus (Kubota et al, 2012). Another marker AspMSD was recently reported for sex identification in different asparagus cultivars and some related species (Mitoma et al, 2018).…”

“…The leaf samples were mechanically homogenized into powder in liquid nitrogen using pestle and mortar. The total genomic DNA was isolated using the plant genomic DNA Kit (Tiangen Biotech, Beijing, China) following a single step DNA extraction method described by Yusuke Ii, Uno, et al (2012). The quantity and quality of the DNA were measured with the absorbance at 260 and 280 nm with a spectrophotometer.…”

“…Approximately, 760 RAPD‐type markers were tested to investigate male‐specific region around the M ‐locus of asparagus; however, they found only one primer pair (OPC15), resulting in two polymorphisms specific to M ‐locus (Jiang & Sink, 1997). Previously, a polymerase chain reaction (PCR)‐based male‐specific DNA marker (Asp1‐T7) was developed using the M ‐locus linked bacterial artificial chromosome (BAC) clones (Ii, Uno, et al, 2012; Jamsari et al, 2004; Kanno et al, 2017). The dominant marker Asp1‐T7 can be used for sex identification in several cultivars from different origins (Nakayama et al, 2006) and was applicable in some related species such as A. maritimus , A. schoberioides and A. kiusianus (Kubota et al, 2012).…”