Plant Propagation. An Overview of Asexual Reproduction

Transcription

1 Plant Propagation Plant Propagation An Overview of Asexual Reproduction Grafting and Budding T or Shield Budding Approach Grafting The Four-Flap Graft Texas Inlay Bark Graft Greenwood Grafting Whip Grafting Collecting and Storing Graftwood Layering Air Layering for Difficult to Root Plants Plant Tissue and Organ Culture (in vitro culture) Micropropagation Aggie Horticulture Extension Horticulture [10/08/ :40:18 a.m.]

2 T or Shield Budding Plant Propagation T or Shield Budding T budding or shield budding is a special grafting technique in which the scion piece is reduced to a single bud. As with other techniques of asexual propagation, the resulting plants are clones (genetically identical plants reproduced from one individual entirely by vegetative means). The plant being propagated (represented by the bud) is referred to as the scion, while the plant being grafted onto is referred to as the rootstock, or simply stock. A small branch with several buds suitable for T budding on it is often called a bud stick. Successful T budding requires that the scion material have fully-formed, mature, dormant buds, and that the rootstock be in a condition of active growth such that the "bark is slipping". This means that the vascular cambium is actively growing, and the bark can be peeled easily from the stock piece with little damage. T budding can be performed on certain fruit trees (peaches, for example) in June using cold stored budsticks and field grown seedling rootstocks. Many deciduous trees are budded in late July or early August after the current seasons buds have developed fully and are dormant using field grown seedlings that have slipping bark. Bud sticks having plump, healthy buds are suitable scions. These budsticks should be on branches that exhibited good growth during the current season, rather than ones from the interior of trees that have slender stems and closely spaced, small buds. Thick water sprouts that grew very vigorously are often poor scions. Leaf blades are clipped from the budsticks, leaving the petiole intact. This leaves a convenient "handle" for holding the bud while it is cut from the budstick. (1 of 4) [10/08/ :40:46 a.m.]

3 T or Shield Budding The bud and a small sliver of the wood underneath it are cut from the budstick using an upward slicing motion. The cut should begin about 1/2 to 3/4 inch below the bud, and should go deep enough into the wood so that when the cut is finished about 1/2 to 3/4 above the bud, the bark and a small sliver of wood are cut off. A perpendicular cut across the top of the upward cut will separate it from the bud stick. Budding knives should be kept very sharp, so that as little damage as possible is done to the bud. Dull knives strip and tear the wood, leaving cuts that do not heal properly. Buds must be cut from the bud stick just prior to grafting, otherwise they will dry out. Some grafters put the bud in their mouth for the time between when it is removed from the stick and when it is grafted in place, but this practice is not recommended. Speed in grafting is a more suitable solution. Some grafters make a downward cut as the second cut to remove the bud from the budstick. This works well as long as it does not result in too much of the underlying wood being removed with the bud. (2 of 4) [10/08/ :40:46 a.m.]

4 T or Shield Budding A vertical cut is made on the stem of the root stock. The cut should be deep enough to insure that the bark will separate at the cambium. The "T is then crossed." That is, a perpendicular cut is made at the upper end of the vertical cut. In areas with heavy rainfall during the grafting season, or in species in which the rootstock is likely to "bleed" heavily, an upside down, or inverted T bud can be used to prevent water or sap from pooling in the graft. The bark is carefully slipped from the stem of the rootstock exposing a "pocket" into which the bud shield can be placed. Care should be taken not to tear the flaps of bark in the process of spreading them. If the bark does not slip easily, this indicates that the stock is not in active growth and the process should be conducted later when active growth has resumed. An alternative method for budding which does not require the bark to slip is the technique of chip budding in which the bud is cut out with a "chip" of the underlying wood. This requires that a chip of corresponding size be cut out of the stock piece in order to align the cambia for proper graft healing. (3 of 4) [10/08/ :40:46 a.m.]

5 T or Shield Budding The bud shield is carefully slipped in between the bark flaps. The top of the bark strip on the bud shield is trimmed to fit tightly against the horizontal cut (the cross of the T) so that the bud fits within the "pocket" snuggly. The bark flaps are held tightly against the bud as they are wrapped with a budding rubber, grafting tape or other suitable closure. This closure must either breakdown by weathering (as budding rubbers do), or must be removed in 2 to 3 weeks after the union has healed. If the material does not break down, it will girdle the rootstock. After the union has healed, the upper part of the rootstock plant can be cut away to force the bud to grow (as would be the case for June budding). If the grafting is done in the late summer, the bud likely will need to overwinter prior to resuming growth. In this case, the upper portion of the rootstock is usually removed during the dormant season, either in late winter or early spring. After the upper portion of the rootstock is removed, the scion bud grows vigorously. (4 of 4) [10/08/ :40:46 a.m.]

6 Approach Grafting Larry Stein Texas Agricultural Extension Service The distinguishing feature of approach grafting is that two independently growing, self-sustaining plants are grafted together. This self-sustaining characteristic of both plants which are to be grafted insures survival of both even if the grafting attempt is, for some reason, not successful. However odds of being successful are greatly enhanced because of the active growing condition of both plants involved and absence of a time limitation required for the healing of the graft union to occur before the dependent scion (top portion) dies from lack of sustenance. The approach grafting procedure is as follows: Plant an adapted, growing plant as close to the base of the non-adapted variety as possible without extensively damaging the root structure of the established plant. From both plants closely position shoots which are at least three-eighths inch diameter and preferably close to the same size. At the point where the union is to occur, a slice of bark one to two inches long is peeled from both stems. The peeled area should be the same size on each. The two peeled surfaces are then bound tightly together with budding or electrical tape. Wrap completely with two complete covers around the area where the two peeled areas are in contact. Remove some of the top portion of the foliage from the adapted variety six to eight inches above the graft union. This will encourage a more rapid healing of the grafted union. The union should be complete in four weeks. This type of grafting is most successful if performed during growth season. After the parts are well united (4 weeks or more),the remainder of the top of the adapted, native variety can be cut off immediately above the graft union and the bottom or root system of the non-adapted, yellowing plant can be cut off immediately below the graft union. The graft union is now completed and the problems of iron chlorosis and indigenous soil pathogens have been solved if the proper rootstock has been used. Immediately after the portion of each plant is removed it may be necessary to reduce the leaf area of the top if wilting occurs because of lack of sufficient root system support. This situation will soon stabilize. If the only problem has been micronutrient (iron chlorosis) deficiency, the top, unadapted variety will not need to be detached from its (1 of 2) [10/08/ :41:05 a.m.]

7 own root system--the approach grafted, adapted variety root system will "feed" the sickly plant what it needs. However, if the purpose of the graft is to control soil borne diseases, the susceptible variety should be detached from its root system and become totally dependent on the root system of the adapted variety. Hypertext markup and graphics colorization by Tammy Kohlleppel and Dan Lineberger. (2 of 2) [10/08/ :41:05 a.m.]

8 The Four-Flap Graft Sammy Helmers and Austin Stockton Texas Agricultural Extension Service Grafting is needed to propagate outstanding varieties of fruits. The four-flap graft is an easy-to-do, successful propagation procedure for amateurs and professional horticulturists. It is ideal for small-caliper trees up to 1 inch in diameter before they are large enough for the Texas-method inlay bark graft. The four-flap grafts most successful when the scion and rootstock pieces are near the same diameter. The best fit is obtained when the scion is slightly larger than the stock. Collecting and Storing Propagation Wood. Collect scion wood from the desired tree while dormant in late February or early March and store until spring propagation time. Optimum scion wood diameter is 3/8 to 3/4 inch; 1-inch diameter is maximum. Select healthy, 1-year-old wood with prominent, well developed buds. Label all propagation wood by cultivar name and place it in a container with moist paper towels. Polyethylene bags make excellent storage containers. Store the wood in a refrigerator at 32 to 36F. Grafting Technique. The best time to graft is April to mid-may when the rootstock is actively growing, the bark "slips" freely and leaflets are 1/2- to 3/4-inch long. Take the scion wood directly form cold storage and use immediately. Do not let the scion sticks dry out during grafting procedures. The following outlines the four-flap grafting method: Use a stock plant with a primary stem or lateral limb of 1/2-to 1-inch diameter. Cut straight across the trunk or limb with sharp pruning shears at the point you wish to graft. If possible, leave one or two side branches below the grafting point, but cut them back to 6 inches. This keeps the tree vigorous, protects it from sunscald and keeps the scion from becoming too tall or whiplike and breaking off. Place the graft 7 to 8 feet above the ground if cattle or horses are grazing near the trees; otherwise place the grafts at a comfortable working height. Roll a rubber band 3 inches down form the top to the stock (figure 1). On the stock plant where the horizontal cut was made, make four vertical, equally spaced cuts 1 1/2 inches long. Make sure the cuts penetrate the bark down to the interior wood (figure 2). (1 of 2) [10/08/ :41:19 a.m.]

9 Hypertext markup and graphics colorization by Gretchen Eagle and Dan Lineberger. (2 of 2) [10/08/ :41:19 a.m.]

10 Texas Inlay Bark Graft George Ray McEachern, Sammy Helmers, Larry Stein and John Lipe Extension Horticulturists Texas Agricultural Extension Service Texas A&M University College Station, TX June 16, 1992 Inlay grafting is one of the best and most popular systems of propagating pecans in Texas. It has been successfully used when other systems have failed because of heat, drought and wind. It has also been successfully used on walnuts, apples, pears, grapes, rabbiteye blueberries and persimmons. The Texas method of inlay grafting, developed by B.G. Sitton, L.D. Romberg, F.R. Brison, B.G. Hancock and others in the 1950s, follows the basic fundamentals of the standard bark graft. However, this technique uses an inlay cut and employs an entirely new system of covering the graft and stock. The inlay occurs when tow parallel cuts are made through the stock bark forming a scion inlay pattern on the stock. Aluminum foil is used as a stock cover, reflecting sunlight and reducing temperatures around the graft. The Foil is covered with polyethylene film to assure constant high relative humidity around the graft. This system not only results in a high percentage of growing grafts, but it is easy to use. All necessary equipment can be carried in an apron since the need for a burdensome wax melter is eliminated. The more stressful the grafting conditions, the more important this grafting technique becomes. This method gained popularity and wide use throughout the pecan industry following numerous method demonstrations and promotions by B.G. Hancock and his many students. Anyone can successfully use the inlay graft by following these instructions and practicing to develop skill in the basic techniques. Figure 1. Use rootstock trunks or major side limbs that are 1 1/2 to 3 1/2 inches in diameter. Leave one or two side branches below the cut to keep the tree vigorous, to protect from sunburn and to keep the graft form overgrowing and blowing out. Cut straight across the trunk or limb with a sharp saw above a straight section of the trunk or limb. Make the cut 7 or 8 feet above ground if cattle or horses are grazing in a native grove. Figure 2. Select a section of stock with a flat surface so the flat cut surface graft stick will fit cambium to cambium without air space separation. Choose a spot on the south or southwestern side so that prevailing winds will blow the graft shoot toward the trunk instead of away from the trunk. If the old bark is rough, cut it down to live bark, forming a clean shield. leave the bark as thick as possible to securely hold the graft. Do not cut through the bark into the wood. Figure 3. Use a knife with a very sharp blade and a sheepfoot point, similar to that (1 of 7) [10/08/ :42:18 a.m.]

11 illustrated. Grafting knife blades are beveled only on one side to give a flat cut. Firmly hold the knife in a closed fist and cut the graft stick with numerous thin slices. Figure 4. The finished graft stick will have one to three buds and three cuts; a slant cut, a long cut and a back cut. The slant cut should begin 1/2 inch below and on the side opposite the lowest bud. It should extend half the distance through the graft stick at approximately a 45 degree angle. The long cut is the same thickness from the slant cut to the end of the graft stick. Make the long cut perfectly flat at the midpoint of the graft stick. The back cut is chisel-shaped and is 1/2 inch long on the back side and lower end. This makes it easier to insert the graft stick and provides additional cambium contact. The long cut can be 1 1/2 to 3 inches long. Figure 5. Place the long cut surface f the graft stick against the clean shield of live bark on the stock. Allow the slant cut to extend above the stock. Firmly hold the graft upright with the left thumb. Begin the first inlay cut at the top of the stock on the right side of the graft stick. Cut through the bark down into the wood. Draw the knife straight down the right side of the graft stick to within 3/4 inch of the bottom portion of the graft stick. It is very important to make this cut straight into the bark. Do not angle the knife to the left or right. Figure 6. Hold the graft firmly in position with the thumb of the right hand. Do not allow the graft to move after the right inlay cut is made. (2 of 7) [10/08/ :42:18 a.m.]

12 Figure 7. Bring the left hand around the back of the stock. Catch the graft with the first three fingers of the left hand and hold in exact position. Make the second inlay cut on the left side to the graft stick, cutting straight into the stock as on the right side. Figure 8. The two parallel inlay cuts through the bark should be exactly the same shape as the long cut section of the graft stick. Figure 9. Peel the bark flap 1/2 inch down between the two parallel inlay cuts. Slide the graft stick between the bark and wood of the stock. There should be no air space between the long cut and the flat wood surface. If the bark does not easily separate from the stock, the cambium is not slipping and you will need to wait several days, then try again. Figure 10. As the graft stick is inserted, press the bark flap against the graft stick with the thumb of the right hand to firmly hold the graft stick in the slot. Apply firm but gentle pressure on top of the graft until it is forced into the inlay slot. Figure 11. Stop pushing the graft stick when the bottom of the slant cut touches the top of the stock. This exposed slant cut surface will form callus and new tissue, which will cover the top to the stock and securely anchor the graft to the stock in 1 to 3 years. Do not push the (3 of 7) [10/08/ :42:18 a.m.]

13 slant cut below the top to the stock because it will separate the graft stick from the flat wood. Figure 12. The graft can be secured by any one of several methods. Eighteen gauge 3/4-inch nails, 5/8-inch flat point staples in a vertical position, budding tape or flagging tape have all been successfully used. Figure 13. Take a 2-inch square of household aluminum foil and tear a line halfway down to the center of the square. Fold the aluminum foil around the stock so that the bottom of the tear fits right under the lowest bud. Figure 14. Fold each side of the divided end of the square of aluminum foil. Cover all cut surfaces with the foil, including the slant cut of the graft stick. Crimp the foil to form a loose mod around the stock. All cut surfaces of the trunk and graft stick should be covered. Figure 15. Cut off one corner of a pint- or quart-sized polyethylene bag. Slip the bag over the graft stick and gently pull it down until the cut corner rests below the lowest bud and above the slant cut. (4 of 7) [10/08/ :42:18 a.m.]

14 Figure 16. Tie the polyethylene bag at the cut corner around the graft just below the lowest bud and above the slant cut so that no air leaks occur. Tie with one wrap of a rubber band, small rubber strip or polyethylene tape so that the graft will not be girdled as it grows. Figure 17. Tie the lower end of the polyethylene bag around the stock with foil covering all of the enclosed area. Make a small puncture above the lower tie to allow water to drain out of the bag. Figure 18. Coat the cut surface of the tip end of the graft stick with orange shellac or white glue. (5 of 7) [10/08/ :42:18 a.m.]

15 Figure 19. The buds on the graft stick should begin groth in 6 weeks. Remove the polyethylene bag and foil when the shoots are over 6 inches long. Keep these shoots pruned back to only 24 inches to prevent wind blowouts. If maximum growth is needed, in 6 to 10 weeks select the strongest shoot and tie it to a brace to prevent it from blowing out. After one year, select the strongest shoot on the graft stick and remove all others. After 2 or 3 years, when three-fourths or all of the trunk is covered with overgrowth, remove all shoots below the graft. Hypertext markup and graphics colorization by Gretchen Eagle and Dan Lineberger. (6 of 7) [10/08/ :42:18 a.m.]

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17 Green Wood Grafting An educational resource of the Texas Agricultural Extension Service. Green Wood Grafting A scripted slide set prepared by Loy Shreve, Jerry Parsons and Sammy Helmers Extension Specialists, Department of Horticultural Sciences Revised August, 1998 by Larry Stein Texas A&M University and Texas Agricultural Extension Service Greenwood grafting allows growers to propagate desired fruit and nut varieties during the growing season after the time for grafting with dormant scions has ended. This method has been successful with pecans, walnuts, apples, apricots and plums; the procedure will probably work with most other deciduous trees and shrubs also. This grafting method can be used from mid-may through September in Texas. In addition to stocks and scions, a sharp knife with a stockman's blade, strips of rubber innertube or polyethylene budding tape and polyethylene squares are needed. The stock should be at least 1 inch in diameter at the point of union. Scion-wood must be the current season's growth. Scions can be stored in an ice chest for up to 5 days without damage if they are kept moist and cool. Place a bag of ice cubes in one corner of the chest to keep the scions in good condition. Use of this Slide Set Advance through the slide presentation by clicking on the "next" link. The images on the individual pages are "clickable", that is, you can see a full screen image when you click on it Propagation Index Go to Index Next [10/08/ :42:45 a.m.]

18 Whip Grafting Whip Grafting Bluefford G. Hancock, George Ray McEachern, and Larry A. Stein Texas Agricultural Extension Service WHIP GRAFTING (also called splice or tongue grafting) is one of the oldest methods of asexual plant propagation known. It is the predominant propagation method used on apples and is widely used on pear. Although most grapes are grown from cuttings in this country, whip grafting is the standard when they are propagated. Whip grafting has been the primary method employed in propagating pecan nursery stock in the southeastern United States. This technique is also used to some extent in the Southeast and west to Louisiana for top-working larger pecan trees on the above-ground portions. Since successful whip grafting is closely correlated to the presence of high humidity, this method has not been used widely in the drier sections of Oklahoma, New Mexico and Texas. A major strong point for whip grafting nursery stock is the smooth and straight trees that are produced by this method. Step 1 Step 2 Step 3 Step 4 Step 5 Step 6 (1 of 2) [10/08/ :43:11 a.m.]

19 Whip Grafting Step 7 Step 8 Step 9 Step 10 Layout, graphics colorization, and html markup by Christine MacAlpine and R. Daniel Lineberger Propagation Index Aggie Horticulture (2 of 2) [10/08/ :43:11 a.m.]

20 Collecting and Storing Graftwood Collecting and Storing Graftwood George Ray McEachern Extension Horticulturist Texas Agricultural Extension Service Texas A&M University College Station, TX June 17, 1992 Successful in-lay and four-flap grafting of pecans is dependent on the availability of suitable graftwood of the desired variety. The wood must be collected in late winter during the dormant season and properly stored until spring grafting time. Collection Collect the graftwood in late February to early March while the tree is still dormant. If the buds have begun to swell and grow, the wood is inferior and cannot be used successfully. Select parent trees of the desired variety that are free of obscure scale, rosette, and disease. Young vigorous trees produce abundant, smooth and large-sized, current-season wood. Moderate-sized trees making normal growth usually have good graftwood in their uppermost limbs. Older trees can be cut back to force vigorous new growth satisfactory for graft wood. Stick preparation Select straight, smooth graftwood from 1-year-old wood 1/4- to 1/2-inch diameter. Cut shoots for grafts into 6-, 12-, or 18-inch lengths to give one, two, or three graft sticks. Figure 1 is graftwood 3/8 inch diameter collected in February for storage as 6, 12, or 18 inch sticks. Each graft stick should contain at least three buds or nodes. Seal the end of the graft sticks with melted wax, grafting paint, or orange shellac. Only 1/4-inch of the end of each stick needs to be treated. When the seal is dry, tie the graft sticks in bundles no more than six each. Label each bundle with permanent ink on a wood or metal write-on label. The variety and year should be recorded on the label. Figure 2 is a graftwood bundle labeled and ready for packing material and storage. (1 of 2) [10/08/ :43:48 a.m.]

21 Collecting and Storing Graftwood Packing material Paper towels, sphagnum moss, or wood shavings can be used as packing material to prevent the graft sticks form drying out. Moist paper towels that have had all of the water squeezed out can be wrapped around each bundle. A second method involves placing the graft stick bundles in slightly moist sphagnum or wood shavings. The wrapped bundles are then placed in polyethylene bags. Plastic bags do not breathe and should not be used. Storage Refrigerate the bags of graft stick bundles at a temperature of 30 to 45 degrees F. Do not allow graftwood to dry out during storage. Take the desired wood out of refrigeration only as needed. Wood should not be heated and recooled during the grafting season. Grafting Trees will be ready for in-lay or four-flap grafting in the spring as the trees begin to grow. The bark should slip during this period. Keep the graftwood in a cool, moist place while grafting. Hypertext markup and graphics colorization by Gretchen Eagle and Dan Lineberger. (2 of 2) [10/08/ :43:48 a.m.]

22 AIR LAYERING FOR DIFFICULT-TO-ROOT PLANTS Everett E. Janne Extension landscape horticulturist Air layering is a useful method of producing roots on the stem of indoor landscape plants that have become "leggy" through the loss of their lower foliage. This method, believed to have been developed centuries ago by the Chinese, has been used successfully as a mean of propagating some of the more difficult-to-root plants. Because it required excessive care and patience, air layering was used only by the highly trained plantsman. The procedure was to wound the stem or branch of a plant and enclose the wounded stem with moist sphagnum moss or similar rooting medium until roots develop from the wounded area. Success was dependent upon the ability of the propagator to keep the rooting medium moist until the roots were formed and large enough to support the new plant. Only since the development of polyethylene film has air layering become a practical method of propagation for the home gardener and amateur horticulturist. Air layering seldom is used on plants that root easily by other less complicated methods, but it is useful for rooting ornamental plants such as ornamental figs, dieffenbachia, croton and others of a herbaceous nature. Woody plants frequently propagated in this manner include magnolia, holly, camelia, azalea and many of the fruit and nut bearing plants such as citrus, apple, pears and pecans. For optimum rooting make air layers in the spring on shoots produced during the previous season or in mid-summer on mature shoots from the current season's growth. On woody plants, stems of pencil size or larger are best. The stem may be much thicker on the more herbaceous plants. Steps for making a successful air layer are illustrated in the following drawings: Figure 1. Method of wounding woody plants such as magnolia, gardenia, rose, fig and similar plants. With a sharp knife, make two parallel cuts about 1 1/2 inches apart around the stem and through the bark and cambium layer. Connect the two parallel cuts with one long cut (a) and remove the ring of bark (b), leaving the inner woody tissue exposed (c). (1 of 5) [10/08/ :44:36 a.m.]

23 Figure 2. Method of wounding plants having less woody stems in preparation for air layering. This method usually is used on foliage plants such as the rubber plant, (Ficus benjamini and Ficus elastica) and the dieffenbachia. (a) With a sharp knife, make a long upward cut from 1 1/2 to 2 inches long, almost to the center of the stem. (b) Insert a wood sliver, toothpick or twisted piece of sphagnum moss into the wound to hold it open and prevent cut tissue from reuniting. At this point, the wounded area may be dusted with one of the commercial rooting compounds to speed up the rooting process. Such compounds, however, do not insure root production on difficult-to-root varieties. Figure 3. Apply a handful of damp sphagnum moss so that it envelopes the wounded portion of the stem. Tying the moss in place with string helps keep it in position while completing the process. The sphagnum moss hould be soaked several hours to insure that it is thoroughly moist. Squeeze out surplus water before using, since excessive moisture will result in decay and deterioration of the plant tissue. (2 of 5) [10/08/ :44:36 a.m.]

24 Figure 4. Using a sheet of polyethylene film approximately 6" X 12" or 8" X 12", depending upon the size of the plant stem, wrap the ball of sphagnum moss using the butchers fold (see insert) to secure a tight seal where the two ends of the sheet are joined. Figure 5. Draw the upper end of the film snugly around stem making sure that none of the moss is exposed. Fasten securely with electricians tape, taking care that the tape extends beyond the film and adheres to the stem. Repeat the procedure on the lower end, again making sure there is a snug fit. Moisture must not escape and excess moisture must not enter when watering or syringing the plants. Support the plant with stake or splint to prevent breakage at the wounded area. (3 of 5) [10/08/ :44:36 a.m.]

25 Figure 6. After the new roots have penetrated the moss ball and are visible on all sides, the rooted branch may be removed from the parent plant. The rooting time will vary with plant variety as well as the season in which it is performed. Figure 7. Remove the newly rooted plant from the parent plant with a sharp knife or pruning shears, making the cut just below the ball of moss and roots. (Not illustrated) Carefully remove the polyethylene film. Without disturbing the roots or removing the ball of moss, plant in a container using a good potting mixture or plant in a well-prepared soil bed. (4 of 5) [10/08/ :44:36 a.m.]

26 Figure 8. Placing a polyethylene tent over the newly potted plant for 4 to 8 days until the root system is well established is helpful as it will aid in preventing excessive loss of moisture. Keep the plant under a light shade and avoid direct sunlight until the new root system is well developed. Many plants are lost in the final stage of the process because the root system is not sufficiently developed to sustain the top portion of the new plant. By utilizing the plastic tent illustrated in figure 8 or by keeping the new plant in a humid environment, it is possible to develop a good root system on rather large cuttings. Once the plant is well established, it is best to harden off the foliage by gradually exposing it to normal atmosphere. This can be done by cutting a few holes every few days in the plastic tent to reduce the humidity until it is similar to the external atmosphere. Educational programs conducted by the Texas Agricultural Extension Service serve people of all ages without regard to race, color, sex, disability, religion, age or national origin. Issued in furtherance of Cooperative Extension Work in Agriculture and Home Economics, Acts of Congress of May 8,1914, as amended, and June 30, 1914, in cooperation with the United States Department of Agriculture, Zerle L. Carpenter, Director, Texas Agricultural Extension Service, The Texas A&M University System. Hypertext markup by Tammy Kohlleppel and Dan Lineberger. (5 of 5) [10/08/ :44:36 a.m.]

27 Woody Plant Tissue Culture TISSUE CULTURE OF WOODY PLANTS R. Daniel Lineberger Professor of Horticulture Texas A&M University, College Station, TX Much has been written and more has been said about the applications of plant tissue culture to the nursery industry. The subject is still poorly understood by a majority of plant propagators. Tissue culture is often looked upon as being practical only for propagation of those plants which are difficult to propagate by conventional methods. This notion is quickly being laid to rest. An industry is emerging in various areas throughout the country which is taking advantage of technological advances in the realm of tissue culture of woody plants. This technology suggests that propagation by tissue culture is indeed applicable to the "difficult to propagate" species, and equally as important, it may offer economic advantages for some species which are considered relatively "easy to propagate". The intent of this communication is to briefly describe the contemporary techniques of tissue culture of woody plants, and to review the advantages and limitations of tissue culture as a propagation method. THE METHODOLOGY OF TISSUE CULTURE The variety of techniques which can be used to get plant development in vitro (that is, by techniques such as tissue culture) is considerable and completely dependent upon the species in question. Single cells of leaf tissue can regenerate whole plants, as can shoot tips, leaf pieces, root pieces, lateral buds, or stem sections. Not all of these methods are applicable to woody plants, and indeed, not all have been applied to a commercial level to any plants. Shoot tip culture is the method in widest use for the mass propagation of woody species. An actively growing shoot tip is surface sterilized and placed on a defined culture medium under sterile conditions. The culture medium contains inorganic and organic salts (macronutrients, micronutrients and vitamins) as well as an energy source (sucrose or table sugar), growth regulators, and agar )to gel the medium). If the growth regulators are appropriately balanced, the shoot tip elongates, lateral buds break and begin growth, and adventitious shoots are also produced on the stem piece. This rapid proliferation of shoots results in masses of shoots being produced from a single shoot tip. Up to a hundred shoots may be produced in as little as eight to twelve weeks from a single tip. The number of shoots produced and the rapidity of shoot proliferation varies between species, and in some cases, between cultivars of a single species. Shoots are removed from the cultures at regular intervals and a portion of the mass is replaced on fresh media to continue proliferation. The small shoots which are removed are then rooted in a separate medium, either a sterile gelled medium or a peat-perlite medium (in much the same fashion as conventional woody cuttings). Shoots produced through tissue culture are generally easy to root, even though the same cultivar may be difficult to root by cutting propagation. The resulting rooted shoot is referred to as a "plantlet" because of its miniature size, but these can generally be grown in the greenhouse at a rapid rate and with a high degree of survival. Techniques for rooting tissue cultured shoots are currently receiving a great deal of research attention as are methods for establishing these shoots into the greenhouse environment. (1 of 3) [10/08/ :44:50 a.m.]

28 Woody Plant Tissue Culture THE TISSUE CULTURE LABORATORY Most propagators would probably list the expense of setting up a tissue culture laboratory among the reasons for not getting involved with this technology. A lab does cost more than a mist bed! A more severe limitation is without doubt the absence of personnel skilled in the techniques of tissue culture. An individual who has had prior experience with tissue culture methodology can successfully operate a laboratory with a bare minimum of equipment. An autoclave or sterilizer, an accurate means of weighing chemicals, a small inventory of glassware, and chemical stocks or prepackaged media are all that is required to initiate cultures. The laboratory can be enlarged, more equipment added, and additional personnel hired as the need for such arises. The most critical factors are the ingenuity and experience of the laboratory manager. As a tissue culture laboratory becomes operational, procedures other than the actual culturing process become critical. Maintenance of disease and insect free stock plants with a low titer of bacterial and fungal contamination is critical. Also important is the care and handling of the thousands of plantlets prior to their transfer to the field or containers. The ability to grow off the plantlets ultimately will limit production, rather than any limitation due to the propagation phase. TISSUE CULTURE PROPAGATION OF WOODY SPECIES Major advances in the mass propagation of woody plants have been made over the past ten years. Because of the somewhat variable response of species and cultivars to the in vitro environment, much research was (and still is) needed to define the cultural conditions required by ornamental species. As an example of the variability encountered, we have observed that seedling shoot tip cultures of crabapple produce as few as one or as many as sixty-four shoots per explant after three months in tissue culture. Similar variations in the tendency for multiple shoot production have also been noted with adult clonal material of different apple cultivars. The number of woody plant species which have been clonally propagated through tissue culture is increasing at a rapid rate, and in fact, most commercially important ornamental species have been studied. Perhaps the best publicized of the early research on woody plants involved apples, Douglas fir, and rhododendrons. An extensive commercial scale production program for apple scion cultivars and rootstocks has developed in England and in several European countries. Using the successes of the apple, rhododendron, and maple programs as a basis, many recent advances have been made which will facilitate the mass propagation of other woody species. While it is true that cultivar differences account for lack of success with some plants, a growing body of evidence suggests that taxonomic families behave somewhat alike in tissue culture. A large number of deciduous and evergreen Rhododendron species share common cultural requirements, the most notable of which is the requirement for the same cytokinin for adventitious shoot formation. A similar relationship exists between the trees and shrubs of the Rosaceae (including crabapples, pears, plums, and hawthorns). Most rosaceous species will likely be mass propagated on media very similar to that found to be optimal for apples. The anomalies alluded to by the crabapple seedling responses to tissue culture and the emerging similarities between the requirements of rosaceous trees and shrubs demonstrate that much research is needed to define the conditions necessary for commercial scale tissue culture production programs. As is (2 of 3) [10/08/ :44:50 a.m.]

29 Woody Plant Tissue Culture often found to be the case with propagation systems, those systems described as optimal in the laboratory would likely need modification to be considered economical by the industry. IMPACT OF THE TISSUE CULTURE INDUSTRY As is always the case for any emerging technology, the strongest supporters for the adoption of that technology are likely those involved in its development. Having acknowledged these biases, one must now consider the advantages and limitations of tissue culture propagation, and what its economic impact on the nursery industry will be. Not every nursery will OPERATE a tissue culture laboratory. The trend toward specialization of nursery operations as either "propagators" or "producers" is likely to continue in the future. Operation and management of a tissue culture facility requires personnel and physical facilities which are quite different from those required by a container or field production nursery. Previously in this report where the establishment of the laboratory was considered, the relatively small initial investment in physical facilities and equipment was discussed. It should be reemphasized at this point, that the training and quality of the laboratory manager is more important than the laboratory itself, and that this expertise is not currently available among contemporary nursery propagators. Not every nursery will NEED a tissue culture laboratory. Commercial operations specializing in mass propagation of nursery crops can likely produce a sufficient number of plants to satisfy the market. As with any other technology, the success of tissue culture within the nursery industry will be determined by economic factors. Can the plantlets be produced in sufficient numbers as a cost to compare with traditional propagation? Will the propagules fit into the established production sequence in the nursery industry? Are the propagules true-to-type? These answers must be obtained on a species by species, cultivar by cultivar basis. (3 of 3) [10/08/ :44:50 a.m.]

30 Whip Grafting Step 1. Seedling rootstocks, 1 to 2 years of age, are usually used for whip grafting purposes. The diameter at the upper portion of the root generally ranges from 3/8 to 3/4 inch, however, stocks up to 1 inch in diameter can be used. The season for whip grafting is February to early or mid March in most areas, or while the stock is dormant. Whip Grafting Index Propagation Index Aggie Horticulture [10/08/ :45:22 a.m.]

31 Whip Grafting Step 2. Select 1-year graft or scion wood in the dormant season. Size should correspond to size of available root stocks. Each graft stick should contain at least two sets of buds. Graftwood should be selected and stored in late January. A knife with a thin blade shaped as shown above and made from high-quality steel is desirable for whip grafting. Make sure that the knife will take a fine edge and hold it under a heavy work load. Whip Grafting Index Propagation Index Aggie Horticulture [10/08/ :45:35 a.m.]

32 Whip Grafting Step 3. Expose about 4 to 6 inches of the upper portion of the tap root. Where only a few trees are involved, the entire process of soil removal may be accomplished by the use of a hoe. However, if the graft is secured with poly tape, the graft can be placed above the soil line as well. Grafter is poised to make initial slanting cut on rootstock. Note the slight angle of the knife blade. The stock of the seedling rootstock fits into the groove or notch formed by the thumb and forefinger of the knife-hand. This serves to stabilize the stock and to provide a guide as the cut is made. Whip Grafting Index Propagation Index Aggie Horticulture [10/08/ :45:46 a.m.]

33 Whip Grafting Step 4. Pull the knife upward with the blade angled about 45 degrees, making a smooth and straight diagonal cut. This slanting straight-plane cut should be 2 to 3 inches long. Try to make this cut with one stroke of the knife. Whip Grafting Index Propagation Index Aggie Horticulture [10/08/ :46:08 a.m.]

34 Whip Grafting Step 5. Place the knife at a spot on the slant cut approximately one third of the distance from the tip to the heel (or bottom) of the cut. Make a "tongue" cut by working the knife blade downward for a distance of 1 to 1 1/2 inches. Take care to prevent splitting the stock. Use fore-finger of the left hand to brace the stock. Note in the inset that the cut is neither parallel to the grain of the stock nor to the slanting cut, but is actually between the two. Whip Grafting Index Propagation Index Aggie Horticulture [10/08/ :46:28 a.m.]

35 Whip Grafting Step 6. Hold the scion wood securely in the left hand, but with care to prevent injury to the buds. Place knife at an angle to the scion and make a slanting cut (see inset) by pushing the blade away from the body. This straight-plane cut should be made as similar to the cut on the rootstock as possible. Whip Grafting Index Propagation Index Aggie Horticulture [10/08/ :46:45 a.m.]

36 Whip Grafting Step 7. Make the "tongue" cut on the scion by placing the knife blade at a point about one-third of the distance down from the tip. Pull the blade downward at an angle that is about halfway between the grain of the scion and plane of the slant cut. (See inset.) Note that the thumb of the knife hand serves as a guide for a controlled cut, while the forefinger of the left hand stabilizes the scion. Whip Grafting Index Propagation Index Aggie Horticulture [10/08/ :46:59 a.m.]

37 Whip Grafting Step 8. Slip the plane cut surface of the scion down to the slant cut of the stock until the two "tongue" cuts mesh together. The cambium layers of the stock and scion must be aligned if a union is to be obtained. An uneven or wavy cut will result in gaps between the two surfaces. If the two cuts are made properly, the stock and scion will appear to be one. (See inset.) Whip Grafting Index Propagation Index Aggie Horticulture [10/08/ :47:14 a.m.]

38 Whip Grafting Step 9. Wrap the graft securely with masking tape or a special grafting tape. Polyethylene budding tape may be used for this wrap, but may require cutting at a later date to prevent girdling. Make certain that the cambium layers of the scion and stock remain aligned during the wrapping process. Note in the inset that the wrap extends from below the graft union to a point slightly above. This is essential to prevent drying of the cut surfaces before callousing takes place. Whip Grafting Index Propagation Index Aggie Horticulture [10/08/ :47:23 a.m.]

39 Whip Grafting Step 10. Firm moist topsoil around the whip graft to prevent drying. Ideally, the soil should cover all of the taped area with the lower bud group on the scion exposed. The cut surface at the top of the scion stick may be coated with orange shellac or with wax to prevent excessive drying. Of course, if the graft was placed above the soil line, this step would not be necessary. Remember, regardless if the graft is placed above or below the soil, the tape which originally secured the graft must be removed. This can be done as late as one year, but preferably after 3 to 4 months. Whip Grafting Index Propagation Index Aggie Horticulture [10/08/ :47:35 a.m.]