Principles of Plant Genetics and Breeding. George Acquaah

      Plants growing in the greenhouse should be provided the proper intensity and duration of light. If the species is photoperiod sensitive, the lighting should be adjusted accordingly. Proper temperature is required for proper plant growth and development. In some species, a special temperature treatment (vernalization, usually some period of low temperature) is required for flower induction. Furthermore, temperature affects pollen shed in flowers. Consequently, extreme temperatures may cause inadequate amounts of pollen to be shed for successful artificial pollination. Pollen quantity and quality are influenced by the relative humidity of the growing environment. Extreme moisture conditions should be avoided.

      6.5.2 Synchronization of flowering

In artificial pollination, the breeder should be familiar with the species to know its flowering habits regarding time from planting to flowering, duration of flowering, mechanisms and timing of natural anther dehiscence and fertilization, and time of peak pollen production, in order to take advantage of the window of opportunity of anthesis (pollen shed) for best crossing outcomes. To ensure that parents in a crossing program will have flowers at the same time, the practice of staggered planting is recommended: to plant sets of parents at different times. This way, a late‐planted early flowering genotype may be pollinated by an early‐planted late flowering genotype. When depending on natural pollination, interspersed planting on different dates will favor even pollen distribution.

      Photoperiod may be manipulated in photoperiod‐sensitive species to delay or advance flowering as appropriate, in order to synchronize flowering of the parents in a cross. Other techniques that have been used in specific cases include manipulation of temperature and planting density, removal of older flowers to induce new flushes of flowers, and pinching (e.g. removal of plant apex to induce tillering or branching for additional flowers). In corn, the silk of an early flowering inbred parent may be cut back to delay the time to readiness for pollination.

      6.5.3 Selecting female parents and suitable flowers

      After selecting lines to be parents in a cross, it is necessary in artificial crosses to designate one parent as female (as previously stated), as well as identify which type of flowers on the parent would be most desirable to cross. In crossing programs in which the CMS system is being used, it is critical to know which plants to use as females (these would be the male sterile genotypes, or A and B lines; see Chapter 19). Because the pollen or male gamete is practically without cytoplasm, and because certain genes occur in the extranuclear genome (such as CMS), it is critical that parents selected as female plants be selected judiciously.

      Markers are important to plant breeding, as was discussed previously. Some markers may be used to distinguish between selfed and hybrid seed on the female plant. For example, in sorghum, waxy endosperm is conditioned by a recessive allele while normal endosperm is under the control of the dominant allele. If a waxy female is crossed with a normal male, all F₁ seed with waxy endosperm would be products of selfing (undesirable) while normal seed would indicate a successful hybrid. Other markers, molecular and morphological, may be strategically included in a crossing program to allow the authentication of hybridity. In terms of flower characteristics, bigger flowers are easier to handle than tiny ones. Whenever possible, the parent with bigger flowers should be used as female.

      Another critical aspect of flower physiology is the age of the flower when it is most receptive to pollination. The breeder usually determines the optimal stage of flower maturity by examining its physical appearance. Tell‐tale signs depend on species. Usually, fully opened flowers would have already been pollinated by undesirable pollen. In most plant species flowers are emasculated in the bud stage just as the petals begin to show through the bud. Rice is ready in the boot stage, whereas wheat is best emasculated when florets are light green, with well‐developed but still green anthers and feathery stigmas that extend about a quarter of the length of the florets. Furthermore, flowers in the same inflorescence usually have different maturity levels. In species such as the broad bean (Vicia faba), the first inflorescence is more suitable for crossing than later ones. Also, flowers at the base and middle of inflorescences give better results than those at the top. Flowers in the inflorescence that are not used for crossing may be removed, while the ones that are used in crossing may be marked with a label or small clip or peg.

6.6 Emasculation

      The process of making a bisexual flower female by removing the male parts or incapacitating them is called emasculation. It should be pointed out right away that emasculation is not a universal requirement for artificial crossing of plants. Species with fertility‐regulating mechanisms (e.g. male sterility, self‐incompatibility, protogyny, monoecy, dioecy) may be crossed without the often tedious and time‐consuming process of emasculation.

      6.6.1 Factors to consider for success

      Apart from picking the right flowers, it is critical to know the duration of stigma receptivity and pollen viability. The maximum time between emasculation and pollination that can be tolerated varies among species. Since the anthers were removed before they were mature, the female parts are often not yet receptive at the moment of emasculation. This makes it necessary to pollinate at a later time, either during the same day or even later. The caution to observe is that prolonged delay between the two operations increases the chance of contamination from undesirable pollen. To reduce this risk, emasculated flowers may be covered with bags (e.g. glassine, paper, or cloth).

      Pollen quality and quantity varies with the weather and time of day. For example, in chickpea, some breeders prefer to emasculate in the evening and pollinate in the morning. Because emasculation is done before anthers are mature in species such as wheat and barley, pollination is done two to three days later, when the stigma is receptive. In extreme cases, such as in sugar beet, pollination may immediately follow emasculation or be delayed for up to 12 days.

      6.6.2 Methods of emasculation

      There are several techniques of emasculation used by plant breeders that include the use of instruments or chemicals. A pair of forceps or tweezers is one of the most widely used instruments in the emasculation of flowers. Different shapes and sizes are used according to the size and structure of the flower. The methods of emasculation may be classified as direct or indirect.

       Direct anther emasculation

      The technique of removing anthers from selected flowers is the most common procedure for emasculation of flowers (usually using a pair of forceps). When handling plants with inflorescence, it is important to first thin out the bunch by removing immature flowers as well as old ones. This will improve the survival of the emasculated flowers. Breeders of various crops have developed convenient ways of removing the anthers. Sometimes, the sepals are first removed, followed by the petals, before access is gained to the anthers. In soybean and sesame, a skilled person may be able to remove the petals and anthers in one attempt. In flowers such as soybean, the pedicel is easily broken as a result of physical handling of the delicate flower during emasculation. In wheat and barley, the florets are clipped with scissors. Specific techniques for specific crops are discussed in part II of this book.

       Indirect anther emasculation

      In these methods, the anthers are incapacitated without being removed from the flower. Incapacitation is achieved in several ways as follows:

       Thermal inactivationThe inflorescence is first thinned out to leave only flowers at the proper stage for emasculation. It is then immersed in hot water (e.g. held in a thermos bottle) to kill the pollen without injuring the pistil. The temperature and time of immersion is variable (e.g. 43 °C for 5 minutes in rice; 47–48 °C for 10 minutes in sorghum). The inflorescence is allowed to dry before pollinating in about 30–60 minutes.

       Alcohol