Well location.

<p>Well location.</p

Year-Round Reproduction and Induced Spawning of Chinese Amphioxus, <i>Branchiostoma belcheri</i>, in Laboratory

<div><p>Amphioxus is a best candidate for studying the evolutionary and developmental mechanisms of vertebrates, because of its vertebrate-like but much simpler morphology, embryonic development and genome structure. Producing live amphioxus embryos throughout the year is an ideal for comparative evolution and developmental studies. However, all amphioxus species have distinct breeding seasons in the wild and laboratory. We recently found that Chinese amphioxus <i>B</i><i>. belcheri</i> could reproduce repeatedly beyond its natural breeding season when reared under proper conditions. In this study, we were able to extend further and produce embryos throughout the year from October 2011 to October 2012. We found all examined animals had spawned repeatedly during the examined period. In addition, both lancelets <i>B</i><i>. belcheri</i> and <i>B</i><i>. japonicum</i> could be induced to spawn by heat-shock method, although the induced spawning efficiency was not as high as that observed in the European lancelet. In general, we have succeeded in producing <i>B</i><i>. belcheri</i> embryos almost daily throughout the year. This advancement will provide essential embryonic material for evolutionary and developmental studies, and have great implications for the cultivation and spawning induction of other amphioxus species.</p> </div

Year-round spawning inductions in <i>B</i><i>. belcheri</i> from 25^th July 2011 to 23^rd October 2012.

<p>Percentage is the number of spawned animals (males or females count separately) over the total number of temperature shocked males or females in each examined date. Inductions using temperature shift from 19°C to 27°C (Method I, from 25^th July 2011 to 12^th April 2012) and from 22°C to 27°C (Method II, from 13^th April to 23^rd October 2012) are shown in the figures <b>a</b> and <b>b</b>. The Figure <b>c</b> shows the cumulative spawning percentages of males and females using Method I or II. Statistical analyses of the data were carried out using SPASS software (version 16.0) and two-tailed chi-squared test. All raw induction records are listed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0075461#pone.0075461.s004" target="_blank">Table S3</a>.</p

Pressure distribution in the reservoir considering both matrix shrinkage and cleat deformation.

<p>Pressure distribution in the reservoir considering both matrix shrinkage and cleat deformation.</p

Comparison of daily gas rate.

<p>Comparison of daily gas rate.</p

Permeability change curve at point 1.

<p>Permeability change curve at point 1.</p

Pressure change curve at point 1.

<p>Pressure change curve at point 1.</p

Percentage of induced spawning dates of <i>B</i><i>. belcheri</i> for each of 12 examined months.

<p>For each month, the spawning percentage is the number of spawning dates (males only, females only, or both sexes) over the total number of dates on which the spawning induction are conducted. The periods used the two-shock systems are respectively highlighted by black and grey horizontal bars. Natural breeding of <i>B</i><i>. belcheri</i> occurs from June to September.</p

Permeability models used in the simulation.

<p>Permeability models used in the simulation.</p

Comparison of cumulative production.

<p>Comparison of cumulative production.</p