VEGETATIVE PROPAGATION OF BLUE SPRUCE (Picea pungens Engelm.) BY STEM CUTTINGS

BY Anne M. Wagner

A Thesis submitted to the Graduate School

in partial fulfillment of the requirements

for the Degree

. M aster of Science

Major Subject: Horticulture

New Mexico' State University

Las Cruces, New Mexico

August 19'88.

Propagation of Blue Spruce (Picea pungens EngcIm.) by Stem ,,:,tt.nac:"

a thesis prepared by Anne M. Wagner in partial fulfillment of the

,.....'rn"·"'t5 for the degree, Master of Science, has been approved and accepted by

H. Matchett of the Graduate School

1988

Dr. Jamcs T. Fisher, Chairman Dr. Dennis Clason Dr. John G. Mexal Dr. Mary A. O'Conncll

n

ACKNOWLEDGMENTS

I would like to take this opportunity to thank Dr. Jim Fisher who gave me the opportunity and support to do this research.

Thanks to the members of my

committee, Drs. John Mexal, Mary O'Connell and Dennis Clason, for their advice "and encouragement. Many thanks to Dr. Leigh Murray who spent countless hours aiding in the design and analysis of this study. In addition, I'd like to acknowledge the help and input of Greg Fancher without whom this project would have been more difficult, if not impossible. I'd also like to thank everyone at the Mora Ke~)ealrcn

Center for their help. And to Paul Schaeffer, fellow graduate student, my

....,QJ"'...... for helping me out and giving me moral support along the way ..

Many thanks to my family and friends for their encouragement and faith in me, my parents for their emotional and fmancial support. Finally, I would to thank John Harrington, for his advice, support, his expertise in graphics, but for keeping me sane these last few months.

ill

,

VITA

March 5, 1959 - Born at Concordia, Kansas

1981 - B.S., Fort Hays State University,Hays, Kansas

1981 - 1983 - Forestry Extensionist, U. S!Peace Corps, Ecuador

1986 -1988 - Teaching Assistant, Department of Agronomy and Horticulture, New Mexico State University

PROFESSIONAL AND HONORARY SOCIETIES

tional Society QfTropical Foresters

ABSTRACT

VEGETATIVE PROPAGATION OF BLUE SPRUCE (Picea pllllgens Engelm.) BY STEM CUTTINGS

BY ANNE M. WAGNER

Master of Science in Horticulture New Mexico State University Las Cruces, New Mexico, 1988 Dr. James T. Fisher, Chairman

Techniques for vegetative propagation of blue spruce (Picea pungens EngelrU.) stem cuttings were investigated. Time of collection 'of the cuttings, application exogenous rooting hormones and source differences were examined. In apdition, . position, cutting length, caliper and fresh weight were'looked at in relation rooting response.

v

Using 100year-old field-grown blue spruce at the Mora Research Center, terminal cuttings were taken from 120 trees every 4 weeks from December 1986 to May 1987; Three New Mexico sources were used. From each source, 10 trees were assigned to each of 4 hormone treatment levels.

The hormone used was

3-indole-butyric acid (IBA) and was applied as a 5 sec. quick dip to the basal end of the cutting. Hormone levels were control (no IBA), 2500 ppm IBA, 5000 ppm IBA and 10,000 ppm IBA. Cuttings were placed under a wet tent with bottom heat of 20°C for 20 weeks.

Rooting, shoot activity and root characteristics were

quantified.

Overall, rooting was low, with 107 of 1440 rooting.

In addition to higher

rooting rates for cuttings taken in December and February, number of roots initiated, root length and root weight were highest for cuttings made in December. Hormone level was significant only in rooting response. The levels most favorable for rooting were the control and 2500 ppm IBA. Source differences were· also seen in rooting response. Cuttings from the Cloudcroft source rooted at higher levels than did the other 2 sources. The cuttings from the lower part of the tree were more likely to root, have more primaries initiated and more likely to break bud. Shorter cuttings were more likely to initiate. roots than longer cuttings.

From this study, it appears as if timing of collection is the variable of greatest importance in rooting of blue spruce stem cuttings. The most favorable time of collection for root initiation, root biomass, number of roots initiated and root length was December. Hormone treatment did not appear to enhance rooting or alter rooting characteristics, except in the later collections. Cutting position and

vi

length appear to play some role in root initiation. activity in root initiation is also discussed.

vii

The possible role of shoot

TABLE OF CONTENTS

LIST OF TABLES .......................................... x

LIST OF FIGURES ........................................ xi

INTRODUCTION .......................................... 1

LITERATURE REVIEW ..................................... 3

Root initiation ............................................. 4

Clonal variation ...........................................

~

6

Juvenility ................................................. 6

Date of collection ........................................... 9

Rooting environment

................................ /. . . . .. 11

MATERIALS AND METHODS .............................. 17

, RESULTS

............................................... 24

Rooting response .......................................... 24

Root analysis .............................•............... 31

Root fresh weight

.......................... :............. 31

Primary root number ...............................•....

~.

36

Root length ...................... '. . . . . . . . . . . . .. . . . . . . . . .. 39

Shoot activity ............................................. 42

Shoot analysis

................................... ~ . . . . . . . . 44

Cutting length ........................................... 47

Final cutting weight ....................................... 5.0

>

DISCUSSION ............................................ 55

Collection date .....' .............. ;........................ 56

vw

Hormone level ............................................ 57

Cutting characteristics

...................................... 57

Root production ........................................... 58

Shoot activity ............................................. 60

Final shoot characteristics .................................... 61

Conclusions .............................................. 61

LITERATURE CITED ..................................... 63

APPENDIXES

A. Weather data for the Mora Research Center ................... 67

B. Propagation bench environmental data

ix

....................... 73

LIST OF TABLES

Table I. Significant interactions .............................................................................. 25

Table 2.. Observed frequencies of rooting by height ratio ...................................... 30

Table 3. Mean root fresh weight by collection date ............................................... 34

. Table 4.. Mean total number of primary roots by collection date ......................... 38

Table 5. Mean sum of root length by collection date ............................................ 41

Table 6. Mean final cutting length by collection date ............................................ 48

Mean fmal shoot fresh weight by collection date .................................... 52

x

LIST OF FIGURES

Figure 1. Percent of cuttings rooted by collection date ........................................ 26

Figure 2. Percent of cuttings rooted by hormone level .......................................... 27

Figure 3. Percent of cuttings rooted by source ...................................................... 29

Figure 4. Logistic regression of initial cutting length and the

pro ba bility 0 f ro 0 ting .......... ............. ............................................................ .... 32

Figure 5. Means of root fresh weight of rooted cuttings by hormone

level and collection date ;.................................................................................. 35

Figure 6 (a - c). Mean root fresh weight of rooted cuttings by

collection date .............:..................................................................................... 37

Figure 7 (a - c). Mean total number ofpriroary roots of rooted

cuttings by collection date ................................................................................ 40

Figure 8 (a - c). Mean sums of root length of rooted cuttings by

collection date ................................ :.................................................................. 43

9. Probability of budbteak of cuttings after harvest

by source ........................................................................................................... 45

10. Probability of bud break of cuttings after harvest

by collection date and hormone level ........ ;..................................................... 46

11. Mean final length of cuttings by source .

and collection date .............. ;.............................................................................. 49

12. Mean final fresh weight of cuttings by source

and collection date ............................................................... f ............................ 53

13. Mean final fresh weight of cuttings by hormone

level and collection date .................................................................................... 54

Xl

INTRODUCTION

Blue spruce (Picea pungens Engelm.)

IS

utilized throughout the United

States primarily as an ornamental but also as a Christmas tree.

The blue

spruce's naturally attractive form and broad ecological adaptiveness has made it a valuable ornamental. The natural range of blue spruce extends through

the southern Rocky Mountains, from southern Idaho to New Mexico. Hanover (1975) identified New Mexico sources as among the best for color development and rapid growth. setting seed until age 30.

Blue spruce matures slowly, generally not

Due to distinct geographic ecotypes exhibiting

variation in form color and growth rates, the ability to vegetatively propagate superior trees would be advantageous in an improvement program.

Variable success has been reported in attempts to vegetatively propagate blue spruce. Thimann and DeLisle (1939, 1942) achicved 80% rooting success with cuttings taken in April from trees 10 to 20 years old, with less success in r

November, and no rooting in other months. rooted

85~/O

Hanover (1975) successfully.

of cuttings from trees 30 to 60 cm tall.

Rooting success of

cuttings from one-year seedlings varied from a low of 10% to a high of 80% (Struve, 1982). Cultivars ·of blue spruce appear to root more rcliably when

cuttings are taken in January than in the summer months (Iseli and Howse, 1981). The consensus among growers appears to be that cuttings should be taken in late winter or early spring and treated with rooting honnones to achieve maximum rooting response.

The objective of this study was to develop methods for vegetatively propagating blue spruce by stem cuttings as well as identifying important factors affecting rooting response. The factors of interest selected were timing of collection and rooting honncine application. In addition, several covariates were examined, which included cutting position on the tree, cutting length and stem caliper of the cuttings.

,

2

LITERATURE REVIEW

. Rooting responses seem to hinge on competence of a cutting to initiate roots. . Rooting in conifers requires the synthesis of root primordia unlike some species which have preformed root initials. Factors which influence the comp.etence of a cutting to initiate roots include:

genotypic differences,

physiological differences and propagation techniques. Three distinct phases of root meristem development have been described cell division, directional growth and cell differentiation.

After the stem cutting is made, callus usually forms.

Callus is

undifferentiated tissue which forms as part of a wound response.

Callus

. develops mainly from cell divisions in the new meristematic area formed in the cortex, after the stem cutting is severed, although phloem and ·cambial cells can also develop callus. After the callus forms, a few of the meristematic cells will form root primordia. Adventitious root meristems (i.e., root primordia) are preceded by lateral extension of callus xylem or by formation and extension of tracheid nests (Cameron and Thomson, 1969).. From the root primordia, roots will initiate and develop.

3

Root initiation

The process of root initiation has been described for cuttings of juvenile Pinus radiata and P. banksiana. The first observable event in root initiation of P. radiata cuttings from seedlings is the formation of a meristematic locus

(Smith and Thorpe, 1975a). Asymmetric division in surrounding meristematic loci leads to the foonation of meristemoids which differentiate into root primordia. Less clear are the events leading to root initiation in P. banksiana. The callus originates from the cortex remaining after stem dieback due to the basal wound (M ontain et al., 1983a, b). The callus tissue differentiates into a complex mass of tissue containing vascular tissue, tracheid nests and resin canals. The root primordia are fooned from the callus apparently associated with the resin canals.

Endogenous factors are known to playa role in root primordia formation as well as root initiation and development. Smith and Thorpe (1975b) found there are two stages when the presence of auxin is essential in root initiation . and development. The first stage is marked by the initial events leading to meristematic locus formation and the second by events immediately preceding meristemoid development.

However, indications are that more than auxins

are needed for root initiation. Hess (1965) proposed that there were 4 rooting co-factors which act with auxin to induce rooting.

Only 2 of the rooting

cofactors were identilled by Hess. The active component of cofactor 3 was identified as isochlorgenic acid and cofactor 4 consisted of oxygenated tcrpenoids. Girouard (1969) found rooting co-factors present in Hedera helix.

4

Girouard further defmed the active components of the rooting cofactors. Cofactor 1 could not be identified. cofactor 2 contained chlorogenic acid" cofactor 3 was found to consist of chlorogenic acid and an unknown promoter as well as isochlorogenic acid, and cofactor 4 was again identified as oxygenated terpenoids. Comparing products of easy-to-root and hard-to-root pear cultivars, Fadl and Hartmann (1967) identified a rooting co-factor which they suggested could be a condensation product between exogenous auxin and an endogenous phenolic compound.

Furthermore, extracts from the

difficult-to-root cultivar lacked this compound.

Haissig,(1974) argued in a review of the effect of auxin on rooting, that while auxin is a key to rooting, another compound needs to be present, ' possibly an auxin-phenol complex. Evidence that auxin alone cannot induce, rooting is suggested by studies with juvenile Pinus banksiana.

When the

tenninal buds were, removed from cuttings, rooting was inhibited and this, condition could not be overcome by exogenous applications of auxin (Haissig. 1982).

Increased rooting responses resulting from the use of N-phenyl

indoyl-3-butyramide or phenyl indole-3-butyrate but not from simple auxin seem to support this theory (Haissig, 1983; 1979). Apparently the synthetic supplemented auxins substitute for the naturally occurring' auxins and cofactors to promote rooting.

5

Clonal variation

Clonal variation is an important factor in rooting potential, yet its underlying cause is not understood. Among trees' from the same seed source, . some will yield more cuttings tHat promptly initiate roots while cuttings from others fail. Because differences in age and physiological state are apparently nonexistent, rootability must be related to genotypic causes. Williams (1987) found between tree variation in mature western white pine which could not be altered by honnone treatment. Donor tree success ranged from a low of 10% to a maximum of 87%. Several species of spruce have been found to vary by tree in response to honnone treatment and bottom heating of rooting medium (Rauter, 1971). It appears as if some internal threshold has to be reached, or a level of competence which is as yet undefmed (Haissig, 1982). This becomes a concern m the selection of individual trees (i.e., specific genotypes) for propagation. Superior individuals or provenances may not root readily and are therefore eliminated from propagation programs. Another related concern is the deselection of genotypes resulting from rooting failure among clones . (Struve, 1982).

Juvenility

A major problem encountered in vegetative propagation of woody plants is ~

the loss of the ability of a cutting to initiate roots. as the stock plant ages. Cuttings from juvenile. trees generally are easier to root than those taken from mature trees.

Adventitious root initiation is considered to be a juvenile

6

characteristic related to ontogeny (Hackett. 1985). Clark (1981) identified four areas linking maturation state and rooting.

The areas of importan
stem anatomy. which causes physical barriers to root initiation, rooting co-factor levels, endogenous rooting inhibitors and the presence of preformed root initials.

Problems associated with maturation are more severe in some clones and cultivars than others (Kester, 1976). However, the end of the juvenile phase of a woody plant is not easily determined. Kester (1976) stated the control of maturation is mainly a function of the development of the vegetative meristem.

The length of the juvenile phase is determined mainly by the

number of cell divisions that have occurred in a vegetative meristem rather than its chronological age. sexual reproduction.

Often maturity is defined as the capacity for

However, while the ability to set seed is rather easily

determined, it appears that is not terms of rooting capacity.

~ecessarily

a good indicator of juvenility in

There is evidence that loss of rooting potential

often is not marked by the arrival of the capacity for sexual reproduction and therefore can be related to other factors. Clark (1981) contends the loss of rooting ability and the capacity to form flowers are distinctly different physiological processes.

Roberts and Moeller (1978) examined rooting pot~ntial of Douglas-fir as related to achieving reproductive maturity and found the decrease in rooting not dependent on cone production or flowering.

The reduced rooting

potential was instead correlated to other factors related to physiological

7

maturity.

Rooting potential of 16-year, cone-producing trees was found to

change little when compared with 12-year trees. By age 16 most of the trees were producing cones. However, rooting potential did change within the trees, with a higher rooting potential in the lower one-third of the crown. Although this change was somewhat tree-dependent, Roberts and Moeller suggested the loss of rooting potential may be localized within a tree.

Another phenomenon associated with agmg

IS

that tissues found at

different locations on the same tree differ in juvenility. Paradoxically, tissues at the top of the tree are vegetatively 'mature' but are the youngest tissues in chronological age. Conversely, the oldest tissues found near the base of the tree tend to be more juvenile (Kester, 1976).

In 16-year Douglas-fIr, the

rooting potential of the upper two-thirds of the tree was found to be significantly less than the lower one-third of the crown (Roberts and Moeller, 1978). This would appear to support the argument that the lower portion of the crown is more juvenile, even though in some trees cone production can be most abundant in the lowest whorl of branches (Roberts and Moeller, 1978). Schwabe (1976) implicated proximity to roots as a factor in juvenility of shoots. Specillcally, juvenility is related to a gibberellin-like factor associated with the roots.

Gibberellin inhibits flowering and the accumulation of f

carbohydrates in the shoots.

Rootability has been related to crown position effects occurring in juvenile seedlings as well as sexually mature trees. Phillion and Mitchell (1984) found that cuttings from the lower two-thirds of IS-month conifer seedlings rooted

8

somewhat better for all the clones they examined.

The effect of crown

position on rootability was most pronounced among clones yielding rootable. cuttings exclusively from the lower one-third of the crown. It was speculated· that because this clone was the tallest, it might be escaping juvenility and

the .

drop in rooting was due to this difference in height, and perhaps greater maturity.

Branch order also plays a significant role in rooting potential. Secondary lateral branches tend to root better than do primaries (Farrar and Grace, . 1942).

However, secondaries exhibit more plagioiropic growth, which is ~

common problem encountered in propagation programs.

However, Miller·

(1982) found in Fraser frr cuttings that differences in rooting responses attributed to secondary and primary shoots were eliminated when exogenous auxins were applied to cuttings harvested from them.

Date of coUection

Seasonal variation in rooting response is a major factor in vegetative· propagation. Season obviously plays ~ role in physiological conditioning of the stock plant which in tum affects the rooting response of the cutting. Lanphear and Meahl (1963) found root-forming capacity of cuttings from two evergreen species was seasonal, in that it peaked in late fall and winter. This relationship could not be altered by the· application of an exogenous root-promoting auxin.

9

Roberts and Moeller (1978) concluded from a study on

Doug1as~frr

that

seasonal periodicity of shoot· development has a controlling influence on rooting potential. The rooting potential of Douglas-frr cuttings was found to be correlated with stage of shoot dormancy.

State of rest (dormancy) was

determined by the speed of budbreak. Dormancy increases from the onset of rest until the bud chilling requirement is met, then begins to decrease until budbreak occurs. Dormancy (state of rest) is often measured by the number of days to budbreak (Kobayashi et aI., 1982). Rooting capacity was lowest when shoot dormancy was greatest. When the chilling requirement of shoot buds was met, in January and February, rooting potential was maximum. At midrest, November, auxin promoted rooting in excised cuttings.

Norway

spruce was found to root best when cuttings were taken in April and May (Girouard, 1975). Rooting response was maximized when cuttings were taken just before or during budbreak.

The second best rooting was October to

November, when bud dormancy was not yet complete.

Similar results have been seen in studies on Fraser frr cuttings (Hinesley . and Blazich, 1984; Struve and Blazich'f 1984). Hardwood cuttings root well in late January and February, after the chilling requirement of buds had been met.

When cuttings are . dormant, neither budbreak nor. rooting can be

initiated. Chilling studies indicate that more chilling is needed for budbreak than for root initiation.

Rooting capacity can drop after February as the

date of budbreak becomes imminent. Because collection date determines total chilling units received, this factor has been shown to be the source of greatest year-to-year variability in the rooting of Fraser fir stem cuttings.

10

Even an easy-to-root species such as Salix appears to have distinct phases during which root initiation will occur. Levels of endogenous indoleacetic acid (lAA) and rooting were correlated, but not enough to conclude that rooting response is governed by hormonal levels (Vieitez and Pena, 1969). Tognoni et al. (1977) identified a substance from Picea glauca cuttings which varied with season and could be positively correlated to rooting responses. The substance was tentatively identified as abscissic acid, a known growth inhibitor. Nanda et al. (1968) concluded that the effectiveness of auxin in promoting rooting differed among species as well as season. More recently, Wise et al. (1985) showed that IBA effects varied with date of collection. Lanphear and Meahl (1963), on the other hand, concluded that positive effects derived from high levels of IBA and periods of high root formation were synergistic in stimulating root initiation.

In addition, no correlation was found between

rooting cofactors found and root-forming capacity.

Rooting environment

Success in rooting of cuttings u,:volves more than selection of quality cutting material. The rooting environment is of importance. The propagation environment

should

optimize

any

biological

potential

for

rooting.

Environmental factors are critical, although some appear to be of greater importance than others.

Factors of importance include soil temperature,

water, both soil and relative humidity, light, air temperature, and media composition.

11

Bottom heat, the heating of the propagation bed or rooting medium to temperatures between 18°C and 30°C, is generally accepted as beneficial to the rooting of cuttings. Ticknor (1969) noted that several conifer species show increased rooting in response to bottom heat, but in other species higher soil temperatures were detrimental. In outdoor tent propagation systems, bottom heat of 22°C proved detrimental to some plants (Pellet et aI., 1983). Bottom heat inhibits the rooting of westem hemlock (Shinn, 1983) yet torulosa juniper can only be propagated using bottom heat of 21 to 240C (Wetherington, 1983). Heating of the soil medium (18 - 21°C) generally improves the rooting response of Douglas-fir cuttings (BheIla and Roberts, 1974; Brix and Barker, 1973). However, Rauter (1971) found that bottom heat was detrimental to . several species of spruce, most notably black spruce. temperature was not reported.

Unfortunately, the

The level and effectiveness of bottom heat

appears to be species dependent.

In 1969 van Elk reported that bottom heat in conifers generally resulted in .increased rooting responses.

In addition to improved rooting, Canadian

. hemlock developed a larger root sy"'stem with bottom heat. However, it was noted that other treatments, such as fungicide applications, could compensate for absence of bottom heat.

In examining the benefits of bottom heat Dykeman (1976) separated rooting into two processes:

root initiation and root elongation.

Root

initiation was defined as primarily cell division, and root elongation as cell .' elongation and differentiation. This approach recognized the possibility that

12

each process can have a different optimum temperature and allowed root initiation and elongation responses to be examined separately.

While not

attempting to defme optimum temperatures for each process, Dykeman reported that temperatures of 30° to 35°C resulted in quicker emergence of root initials and more roots initiated in Chrysanthemum and Forsythia. However, temperatures of 35°C inhibited growth and development of the roots, even causing death of the root within days. Temperatures below 25°C were shown to enhance root growth. In practical terms it would be difficult to determine when to change media temperatures in propagation operations. However, the effects of media temperatures on root initiation and development must be known to allow for better choices in selecting media temperature.

Brix and Barker (1973) examined the effects of bottom heat in relation to

air temperature. Western hemlock cuttings collected in the fall responded to no heating of air or soil medium (bjlt kept frost-free).

Douglas-fir cuttings

taken in November rooted only in a cold air-warm soil (200C) environment .. Cuttings were taken September through March. Cold air-warm soil was most beneficial to the rooting of Douglas-fir through January.

February and

March cuttings did not respond to cold air-warm soil treatments. Cuttings from Douglas-fir never responded well to warm air-warm soil treatments, although cuttings from seedlings did root in a warm air-wann soil environment.

13

In another study using Douglas-ftr, Bhella and Roberts (1974) reported cuttings rooted better with media temperatures of 18° and 26°C than with Woe.

The observed increase in bud respiration at 18° and 26°C was

correlated to an increase in rooting. At lower temperatures the decrease in rooting was possibly linked to slowed metabolism, as indicated by lower bud respiration. Higher soil temperatures also caused an increase in basal callus formation, an important step in root initiation.

In a later study, optimum

rooting temperatures for Douglas-ftr were narrowed to between 18° and 21°C. Also interactions between daylength and media temperatures in the rooting environment influenced rooting (Roberts and Moeller, 1978).

Other factors can interact with bottom heat in influencing rooting, including auxin activity, air temperature and transpirational losses.

Scott

(1972) indicated increased auxin activity at higher temperatures. Because of the key role auxin has in root initiation, the possible enhancement of auxin

. activity with raised temperatures would be beneficial.

On the other hand,

increasing media temperatures will also signillcantly increase air temperature

at the cutting level (Gislerod, 1983). The increased air temperature will ... increase the transpiration loss in the shoot, b~t adverse responses can be minimized or eliminated when fog and/or mist systems are used in conjunction with higher temperatures.

Tissue water status can strongly influence rooting success.

A favorable

environment in terms of relative humidity and ample water is critical for survival of the cutting.

By using mist instead of double glass, Hess (1965)

14

found increased ,rooting as well as number of roots initiated and root length. Relative humidity in the two propagation systems were the same but leaf temperature was significantly lower in cuttings under mist. Mist was effective in cooling the cuttings through evaporation.

In addition, less light was

allowed to reach the glass-grown cuttings because shading was used to decrease temperatures through reduced incoming radiation to the cuttings. The combination of higher temperatures, which led to increased transpiration, and reduced photosynthesis under lower light resulted in lower rooting success.

More recently, Howard (1980) found humidity in the rooting

environment greatly influenced root formation.

High leaf water potential has been correlated to rooting success (Loach, 1977). Water uptake from the medium by cuttings is restricted by a contact resistance between cutting and medium. In addition~ there is astern resistance to water uptake"'which develops within several days of stem insertion into the medium.

Therefore, factors influencing tissue water balance in the· cutting

indirectly influences rooting success. (Grange and Loach, 1983a).

One of the major factors influencing water loss is daily solar radiation. As. radiation increases, transpiration also increases (Hess, 1965; Loach, 1977). Grange and Loach (l983b) found leaf tissue water content of cuttings inversely proportional to daily radiation.

Some cuttings can offset the

resistance to water uptake. and .water loss by water absorbtion directly through the leaf.

Whitcomb etal. (1982) developed a system that reduces

transpirational losses. as well the a,inount of mist that must be applied. The

15

wet tent propagation system provides high humidity with limited misting. With a wet tent, water evaporates from the fabric as temperatures rise and the humidity increases inside the tent.

In conclusion, propagating conifers from stem cuttings is difficult. Many factors can be involved which can directly or indirectly determine success or failure of a cutting to root. Stock plant conditioning is of major importance as is defmed by physiological state of maturity and dormancy. hormones may be necessary to optimize rooting capacity. environment is also of interest in

tha~

Post-severance

appropriate growing conditions are

necessary for root initiation and development.

16

Rooting

MATERIALS AND METHODS

Stock plants were selected from a blue spruce provenance study planted in 1978. at the Mora Research Center. From 19 sources. 3 New Mexico sources were selected for this study. Sources were selected based on mean height and color. The 3 sources selected were from Cloudcroft, Junction La Junta near Mora. and Willow Creek on

th~

Gila National Forest. The objectives· of the

study encompassed several aspects, most of which involved techniques of propagating blue spruce. The primary objective was to determine if there were any differences in timing of collection on rooting success of cuttings. Secondary objectives included quantifYing the effects of rooting hormone and the effect on root initiation and development. and any differences in rooting potential among different seed sources. Tertiary factors of interest were the effect of cutting position. basal stem caliper and cutting fresh weight on rooting. The selection of 3 sources was made in an attempt to identify source variation in rooting response. as well as maximize potential rooting success.

From each source. 40 trees were selected. Within each source, the trees were selected based on form, color and height characteristics. A total of 120 trees were used in this study.

All trees were un sheared and the plot was

thinned to a 1.7 m by 2 m spacing the previous year. No fertilizer had been

17

applied during the previous growing season, but had been unifonnly applied in 1985,

Due to the number of cuttingl; available from each tree, 6 collection dates were selected in an attempt to identifY optimum time of collection. Previous work conducted elsewhere indicated blue spruce roots best in late winter to early spring (Hanover 1975).. Collection dates were selected to permit harvest at 4-week intervals beginning in December 1986. Cuttings were harvested on the following dates:

20~2l

December 1986, 17 January 1987, 14 February

1987, 13 March 1987, 9 April. 1987 and 8 May 1987 (see Appendix A for weather data).

Indolebutyric acid (IBA), a synthetic auxin, was used to determine if rooting potential could be altered by treatment with exogenous auxins. Three hormone levels were selected along with a control (no hormone). .

.

The 4

treatments were control, 2500. ppm IBA, 5000 ppm IBA and 10,000 ppm IBA..

Each tree was randomly assigned to one of the 4 hormone levels initially. The study required 12. cuttings from each tree.Cutting·s harvested from each tree at the assigned intervals received the same hormone treatment throughout the course of the study to eliminate tree-to-tree variation. Two cuttings were taken from each tre.e at each collection time. Cuttings from a total of 10 trees per· source received the same (teatmcnt. For each collection, 240 cuttings were stuck, with 60 cuttings for each hormone level. A total of 1440 cuttings were used in the study.

18

•

At each collection, the cuttings were harvested in the same manner. Only primary lateral shoots with terminal buds were harvested and these included, only the growth produced the previous growing season. Initially, tree height was recorded for each tree before cuttings were taken. As each cutting was taken, the vertical distance between the ground and the point of stem severence was recorded (cutting height). Cuttings \vere subsequently placed in paper bags with labels identifying origin by source and tree. All cuttings from a source were taken before any were treated. The maximum

l~g

time between

cutting and sticking of the cuttings was approximately 4 hours.

In the laboratory, the basal end of the cuttings were recut at a 45° angle, and if old wood was present it was removed at the time of recutting. ' Final cutting length, basal stem caliper and cutting fresh weight were ,recorded. Cutting lengths varied from 3.7 cm to an arbitrary maximum of' 12.5 cm. Needles were not stripped from the base of the cuttings.

Cuttings were then treated with a

5-s~cond

quick dip of the basal 2 cm in

the prescribed treatment. The 3 hormone treatments containing indolebutyric acid (IBA) were dissolved in 50% isopropyl alcohol. Control cuttings were dipped in a. 50% alcohol solution.

After drying for' .10 minutes the basal

portion of the cuttings were dipped in a 1:1 Captan fungicide/talc mixture_ The cuttings were then immediately stuck in' 25 cm3 polyethylene containers (Ray Leach tubes) to a depth of approximately 2.5 cm.

19

The containers were previously filled with a 1: 1 venniculite/perlite mix (v/v). The filled trays were placed on the propagation bench in a randomized (by treatment) design.

The trays containing the cuttings were placed in a

preselected zone on the propagation bench to minimize edge effects created by temperature, light and humidity gradients.

Border. cuttings were placed

around the perimeter of the grouped trays to further reduce border effects. The unused zones of the propagation bench were filled with blank tubes with

,

media to prevent heat loss and to maintain uniform bench temperature.

The bench utilized for vegetative propagation is located in the center of the greenhouse with an east-west orientation.

The mist bench system is a

modification of the wet tent system designed by Whitcomb et al.

(1~82).

Fluorescent lights are located 105 em above the floor of the bench to provide 24-hour photoperiod. A Biotherm heating system, used to provide bottom heat, is supported by a redwood frame and covered with expanded metal to accommodate growing containers.

The Biotherm system was installed in

accordance with the manufacturer's specifications. Controls were set at 20°C to provide bottom heat for the cuttings. The sensor which controlled bottom heat was placed in the center of the study.

Relative humidity was kept high through the use of two independent systems applying moisture to the bench. A 100% polyester fabric draped over a pitched metal frame attached to the top of the bench provided the enclosure for maintaining relative humidity. The polyester fabric allowed air to circulate through while keeping the humidity high. Anautomated track-mounted boom

20

located 1.0 m above the bench was controlled by an automatic clock timer. Speeds and frequency were adjusted as needed to maintain humidity 65% 10%.

fan~type

The boom contained nine

particles to the cuttings.

+ /~

nozzles to provide uniform mist

In addition, above the tent was a fog system

controlled by an evaporative leaf moisture meter. The fog system kept the

.

tent wet and helped maintain the humidity in the propagatiori bench. Cuttings were fertilized with Hoaglands solution applied with a hand applicator.

Cuttings were fertilized 3 times a week to compensate for the

effects of leaching from the mist applications.

All environmental data were recorded with a Campbell Scientific Datalogger CR7.

Data recorded included. relative humidity at cutting level

and 1.2 m above the cuttings.

Media temperature was measured with 2

thermocouples; air temperature at cutting level was also measured with 2 thermocouples. Natural light levels were measured with two

LI~200S

silicon

pyranometers, and soil moisture was measured using gypsum blocks in 2 locations (see Appendix B). Soil temperatures were maintained .at 20°C + /~ 2°C, air temperatures were maintained at 20°C + I-5°C. Soil moisture levels were kept at

~0.2

bars + 1- 0.1 bars.

Cuttings were removed from the bench after 20 weeks. Treatment blocks were removed individually, with all measurements being made withIn 72 hours of removal from the bench.

If cuttings were removed in: advance of

evaluation, they were placed in a walk-in refrigerated cooler at 4°C. cuttings were destructively sampled.

21

All

For each cutting the following attributes and measurements were recorded:

Cutting condition:

,

0, dead

l,alive

2, callused

3, rooted

Shoot elongation:

0, no activity

1, budbreak or elongation

Shoot measurements:

shoot length

shoot caliper

shoot green weight

Root measurements: primary root number primary root length secondary root number (total) secondary root length (sum) tertiary root number (total) tertiary root length (sum) total root fresh weight (primary and associated root total) .

22

Primary roots were deflned as originating from the cut end of the cutting, or ""

callus tissue (if present). After evaluation, cuttings and roots were oven-dried at

The experimental design was a split-plot design. design was a 3 ( source) X 4 (hormone) factorial.

The whole plot treatment

Collection date was the split

factor. Cutting height to tree height ratio, [mal cutting length, basal stem caliper and cutting fresh weight were used as covariates. Statistical analyses were done using analysis of variance techniques (GLM, SAS Institute, 1985). were done using the Student-Newman-Keuls test.

Range tests

Discrete data were analyzed

using categorical model analysis (chi-square tests) and logistic regression.

For

analysis, the total number of primary roots was used for each rooted cutting. Root length and root fresh weight were totaled to give a single value for each cutting. • Height ratio was a variable created by taking the ratio of cutting height over the total tree height.

23

RESULTS

Overall rooting was low, with 107 cuttings out of 1440 rooted.

Rooting

response was highest· for the first collection dates, dropping off in the spring. Rooting response varied somewhat with honnone level and source.

Tree-to-tree

variation as well as within-tree variation in rooting response was observed. Table I gives a summary of significant interactions over all variables analyzed.

Rooting response

After restricting cutting condition classification to only two categories, rooted and not rooted, a categorical model analysis (SAS Institute, 1985) was used to test for significant effects. Collection date, honnone level and source showed significant effects on rooting response. Collection date was highly significant with a X 2 value of 6.26 (probability < 0.0001).

By collection date, rooting was highest in

December at 15%, followed by 13% in February. There was a drop in rooting in January; and overall rooting declined steadily after February (Fig. 1). Source and honnone level were significant at the =0.0979, 0.0996, respectively).

10~/Q

level (X 2 = 4.65, 6.26; probability

Best treatments were the control and 2500 ppm

IBA with rooting percentages of 9% and 11%.

Rooting success dropped at the

higher levels of IBA (Fig. 2). Best rooting was found in the Cloudcroft source with

24

Table 1. Significant interactions between variables measured during rooting of IO-year blue spruce stem cuttings from December 1986 to May 1987. SUMMARY OF CATEGORICAL ANALYSES

Source Collection Date

Rooting

Shoot

(:to)

'"

'"

*

Root Root fro wt. no.

'"

Root length

Final Final shoot shoot fro length wt.

*

'*

(*)

Hormone

'"

Source:l: CD

'"

Source * Hormone Hormone

* CD

Horm '" Source

'" >I:

'"

'" *

CD

>I:

'"

SUMMARY OF LOGISTIC REGRESSIONS

Cutting length

*(+)

Ht. ratio

'" * (-)

*(+)

Caliper

*

'"

Fresh wt.

'"

'*

'" = significant, P < 5% (*) = significant, P < 10%, . + ,- = sign of estimated slope

25

PERCENT

Q IV 0­

MONTH

FIGURE 1. PERCENT OF CUTTINGS ROOTED BY COLLECTION DATE. (PERCENT IS BASED ON TOTAL NUMBER OF CUTTINGS TAKEN FOR EACH COLLECTION DATE)

(N

=

240)

.,

PEHCENT

oupp 18/\

FIGURE 2. PERCENT dF CUTTI~GS ROOTED BY HORMONE LEVEL. (PERCENT IS BASED ON TOTAL NUMBER OF CUTTINGS TAKEN FOR EACH COLLECTION DATE (N = 360»

27

rooting of 11 %. The Junction La Junta source and the Willow Creek source had 5% and 7% rooting success, respectively (Fig. 3). However, none of the source by collection, honnone by source, honnone by collection or honnone by source by colled:ion interactions were significant.

In addition, tree-to-tree variation in rooting response of cuttings was observed. Individual tree rooting response varied from 0 to 50%. By source, cuttings from 30 of the 40 trees representing the Cloudcroft source sampled in the study rooted at least once. Cuttings from only 15 of the 40 trees from the Willow Creek source and 14 of 40 trees from the Junction La Junta source rooted at least once.

Logistic regression was used to analyze rooting response in relation to height ratio, initial cutting length. initial caliper and initial green weight.

The natural

logarithm of the ratio of the probability of not rooting over the probability of rooting (called the logit response) was used as a response and the slope and intercept.of the line representing the relationship were calculated. Only height ratio and ·initial cutting length had detectable effects on rooting response. The relation between rooting and height ratio was seen when the height ratio data was transfonned into categorical data (Table 2). Vertical position or"the cutting on the tree did influence rooting success. The chi-square test of independence found that rooting· and cutting height are not independent factors.

The cuttings from the

lower third of the tree showed rooting rates that exceeded the expected value.

28

PEf