U.S. patent number PP20,149 [Application Number 09/539,992] was granted by the patent office on 2009-07-07 for new and distinct somaclonal variety of rose scented geranium.
This patent grant is currently assigned to Council of Scientific and Industrial Research. Invention is credited to Suchitra Banerjee, Suman Preet Singh Khanuja, Sushil Kumar, Gopal Rao Mallavarapu, Laiq ur Rahman, Gauri Saxena, Srikant Sharma.
United States Patent |
PP20,149 |
Saxena , et al. |
July 7, 2009 |
New and distinct somaclonal variety of rose scented geranium
Abstract
A new and distinct somaclone of rose scented geranium
Pelargonium graveolens christened `Parimal` characterized by
distinct morphology and improved oil yield determining
parameters.
Inventors: |
Saxena; Gauri (Lucknow,
IN), Banerjee; Suchitra (Lucknow, IN),
Rahman; Laiq ur (Lucknow, IN), Mallavarapu; Gopal
Rao (Lucknow, IN), Sharma; Srikant (Lucknow,
IN), Khanuja; Suman Preet Singh (Lucknow,
IN), Kumar; Sushil (Lucknow, IN) |
Assignee: |
Council of Scientific and
Industrial Research (New Delhi, IN)
|
Family
ID: |
40811178 |
Appl.
No.: |
09/539,992 |
Filed: |
March 30, 2000 |
Current U.S.
Class: |
PLT/324 |
Current CPC
Class: |
A01H
5/02 (20130101); A01H 6/42 (20180501); Y02A
40/14 (20180101); Y02A 40/10 (20180101) |
Current International
Class: |
A01H
5/00 (20060101) |
Field of
Search: |
;PLT/324 |
References Cited
[Referenced By]
U.S. Patent Documents
Other References
MPEP Chapter 1600--Plant Patents; 11 pages. cited by
examiner.
|
Primary Examiner: Bell; Kent
Attorney, Agent or Firm: Abelman, Frayne & Schwab
Claims
It is claimed:
1. A rose scented Geranium plant named `Parimal`, as herein
illustrated and described.
Description
FIELD
The invention provides a new distinct somaclone of rose scented
geranium P. graveolens christened as `Parimal` characterized by
distinct morphology and improved oil yield determining parameters.
The said novel somaclone has been developed from callus through in
vitro tissue culture technique, without enforced mutagenesis. The
plant possesses characteristic leaf morphology, vigorous growth,
improved oil content per plant, herb yield and oil yield. The oil
produced by this somaclone resembles the oil produced by the
commercial type of bourbon geranium oil in that has equivalent
ratio of citronellol and geraniol.
BACKGROUND OF THE INVENTION
The present invention relates to a new and distinct somaclone of
rose scented geranium Pelargonium graveolens a member of the family
Geraniaceae, which is a stable somaclone of the Indian cultivar
Hemanti. The oil of geranium finds extensive use in perfumery,
cosmetics, food and pharmaceutical industries (Narayana, 1986).
Geranium was introduced in India in the beginning of this century
and since then its cultivation and production of oil remained
restricted to the high altitude regions of Ootacamund, Kodaikanal
and Yercaud areas in South India (Narayana, 1986). India produces
hardly 5 tonnes of geranium oil annually as against its own
requirement of approximate 100 tonnes per year (Narayana 1986). The
production of oil has declined gradually over the years and much of
the requirement is met by imports. This may be due to narrow
genetic base, resulting partially from sexual infertility,
vegetative mode of propagation and restricted area of
cultivation.
In India, two cultivars of geranium are mainly known; these are the
types that produce Algerian or Tunisian quality and the Bourbon or
the Reunion quality of geranium oils; these are given the names
Pelargonium.times.graveolens cv Hemanti and
Pelargonium.times.graveolens cv Bipuli, respectively. The cultivar
Hemanti was found to be more adaptive than the cultivar Bipuli both
in the hilly regions as well as in the unconventional areas of the
plains. On the other hand, the cultivar Bipuli scored better than
the Hemanti mainly because of its better oil yield and improved
organoleptic value of the oil. The perfumery value of geranium oil
is mainly determined by the ratio of citronellol and geraniol and
almost equal contents of these two major terpenoid components is
generally considered to be the characteristic of the best quality
geranium oil.
In an attempt to extend the area of cultivation of geranium to the
Indo-gangetic plains, the better adaptive Hemanti cultivar was
introduced in the Lucknow conditions in India. Subsequently,
efforts were made through different biotechnological methods to
develop clones with high content and desirable composition of the
oil. The calliclone `Parimal `evolved through this invention
represents improvement over the wild type Hemanti parental cultivar
both in terms of quality and quantity determining traits of the
essential oil. This particular somaclone, being consistently
conditions, can offer an improved cultivar for commercial
exploitation.
OBJECTS OF THE INVENTION
The main objective of the inventin is to develop useful somaclonal
variants of rose scented geranium christened as `Parimal `which are
distinct, stable and uniform through successive generations.
Another object is to develop somaclonal variants yielding high
quality and quantity of essential oil.
SUMMARY OF THE INVENTIONS
Accordingly the present invention provides a novel variety of rose
scented geranium called `Parimal `said plant developed through in
vitro tissue culture methods from callus cultures. The invention
further provides a useful somaclonal variant `Parimal`of rose
scented geranium (Pelargonium graveolens cv. Hemanti) yielding high
quality and quantity of essential oil.
DETAILED DESCRIPTIONS
In accordance with the above and other objects, the invention
provides a novel somaclone of rose scented geranium called
`Parimal`. The said plant has been developed though in vitro tissue
cultured on modified Murashige and Skoog's medium (MS), 1962.
The invention further provides a useful somaclonal variant
`Parimal` of rose scented geranium (Pelargonium graveolens cv.
Hemanti) having the following morphological characteristics.
Parentage: Pelargonium graveolens cultivar Hemanti. Propagation:
Exclusively vegetative propograted through terminal stem cuttings.
Time to initiate roots: 15-20 days. Time to produce a rooted young
plant through terminal stem cuttings: 30-35 days. Terminal stem
cuttings: 30-35 days. Root description: Fibrous, brown in color.
Rooting habit: Freely branching, dense. Plant description: General
appearance.--Upright, with larger and dense spreading canopy.
Growth branching habit.--Vigorous, showing extensive basal
tillering. Plant length.--85-95 cm. Growth habit. --Spreading,
round canopy. Plant height, to top of foliar plane.--.about.46 cm.
Stem.--Green, internodes 8-9 cms long, nodes pinkish Main
branches.--Length about 85 cm. diameter about 1.1 cm internode
length about 4 cm. texture pubescent, color 138D, color at nodes
75C. Lateral branches.--Length about 70 cm. diameter about 0.7 cm,
intermode length about 8 cm. texture pubescent, color 138D.
Leaves.--Number per branch 25. Arrangement Alternate. Length
.about.6 cm. Width .about.5 cm. Texture pubescent. Shape thick,
serrated with round lobes. Apex obtuse. Base cordate. Number of
Primary lobes 3. Secondary lobes 12-15. Tip Blunt. Leaf area 20-28
cm.sup.2. Texture (upper surface) pubescent. Texture (lower
surface) pubescent. Petiole .about.8 cm (pinkish). Morphological
marker Less dentated leaf morphotype (Ldl). Leaf/stem ratio (L/S)
2.1. Herb yield/plant 4.5 kgs. % oil content 0.18 (mean). Oil yield
/plant 7.1 gms. (mean). Oil yield (kg/ha) 71.1 (mean). Flower.--No
flowering entire lifetime. Seed.--No seed formation. Disease
Resistance: Exhibited no specific susceptibility towards the
commonly occurring fungus.
This invention further provides the useful somaclonal variant
"Parimal" of rose scented geranium (P. graveolens cv. Hemanti)
having the following essential oil quality determining
characteristics. The average and mean percentage of the respective
constituents of the essential oil are provided hereinbelow:
TABLE-US-00001 Essential oil constituents Percentage (%) Linalool
9.1 Cis + Trans rose oxide 0.60 Menthone 0.2 Isomenthone 7.8
Citronellol 27.6 Geraniol 22.6 Citronellyl formate 5.9 Geranyl
formate 2.1 Citronellyl acetate 0.2 .beta.-caryophyllene 0.2 6,9
guaiadiene 0.6 citronellyl butyrate 1.0 10 Epi-.gamma.-eudesmo 7.3
Citronellyl tiglate 0.8
This invention further provides the said somaclone "Parimal" which
exhibited uniqueness with respect to the following RAPD
primers:MAP-05, MAP-12, MAP-15 and MAP-16.
The methods and examples for the development of the said novel
plant "Parimal" are provided herein below, for illustration of the
invention and should not be construed as limitations on the concept
of the invention. Modifications that may be apparent to those
skilled in the art are deemed to fall within the scope of the
present invention.
BRIEF DESCRIPTION OF THE ACCOMPANYING DRAWINGS
The invention is further illustrated with reference to the
following drawings wherein:
FIG. 1 Field grown plant of somaclone "Parimal" having large round
canopy and profuse branching.
FIG. 2 Field grown plant of wild type parent spreading prostrate
canopy and comparatively less branching.
FIG. 3 Leaves of somaclone "Parimal" and wild type showing
comparatively less dentation in the leaf of the former than that in
the latter.
FIG. (a-d): Variation in the RAPD profiles of wild type parent (C)
with that of the selected somaclone "Parimal" (marked with
(.fwdarw.)arrow); a:primer MAP-5; b:primer MAP-12; c:primer MAP-15;
d: primer MAP-16.
Methods for development of the novel somaclonal variant
"Parimal":
Healthy robust shoots, raised by subculturing the organogenic calli
on modified MS medium with concentrations of BAP and NAA, were
transferred to half strength MS medium with a concentration of
indole butyric acid (UBA) for rhizogenesis. The stem explants were
cultured on modified Murashige and Skoog's medium (MS), 1962
containing the following (in mg/L)-NH.sub.4NO.sub.3 (1650),
KNO.sub.3 (1900), CaCl.sub.2.2H.sub.2O (400), MgSO.sub.4.7H.sub.2O
(370), KH.sub.2PO.sub.4 (170), Na.sub.2EDTA.2H.sub.2O (7.2),
FeSO.sub.4.7H.sub.2O (27.8), MnSO.sub.4.4H.sub.2O (22.3),
ZnSO.sub.4.7H.sub.2O (8.6), H.sub.3BO.sub.3 (6.2), KI (0.83),
Na.sub.2MoO.sub.4.2H.sub.2O (0.25), CuSO.sub.4.5H.sub.2O (0.025),
CoCl.sub.2.6H.sub.2O (0.025), Glycine (2.0), Nicotinic acid (0.5),
Pyrodoxine HCl (0.5) and Thiamine HCl (0.1) with proportion of
naphthelene acetic acid (NAA) and Kinetin (Kn)[2:1]. The best
organogenesis (85%) could be achieved by substituting Kinetin with
6-Benzylaminopurine (BAP).
The rooted plants were initially hardened under glass house
conditions and were subsequently transferred to the field in the
month of October 1996, in replicated trials. Based on the superior
performance for vigorous growth resulting from extensive branching
and enlarged canopy size, comparatively better oil yield over that
of the control wild type cultivar and distinctively improved oil
quality in terms of the major terpenoid constituents, the plant of
this invention (Somaclone "Parimal") was selected for further
observation and evaluations. During the monsoon seasons of 1996-97,
this selected somaclone along with control parent were maintained
under glass-house conditions through stem-cuttings which also
maintained clonal purity. The selected clone "Parimal" was grown in
field in replicated trials along with its parent Hemanti cultivar
for two consecutive winter-summer seasons of 1997-98 and 1998-99,
while during the intermediate monsoon season, they were maintained
under glass-house conditions. The clone "Parimal" maintained its
vigorous growth through extensive branching, better oil content and
improved oil quality over the three consecutive years. The
characteristics of this particular clone (in terms essential oil
quantity and quality determining traits) in relation to that of the
control parent and other clones have been represented in Tables-2
and 3. The results clearly demonstrate the outstanding performance
of clone "Parimal" over that of the control wild type parent and
other selected clones under Lucknow field conditions, indicating
the feasibility of its commercial cultivation in the indo-gangetic
plains. It is pertinent to mention ere that the oil profile of
somaclone "Parimal" is superior than that of the wild type parent
in having equal content of citronellol and geraniol which is a
critical characteristic of best quality commercial geranium oil.
The field grown plant of "Parimal" having large round canopy is
shown in FIG. 1 and FIG. 2 depicts the wild type parent. The leaves
of "Parimal" are as shown in FIG. 3.
Evidences of uniformity and stability
The somaclone "Parimal" has remained stable and uniform for its
morphological characters and showed consistency in performance for
various oil yield and quality determining attributes during its
field evaluation under Lucknow conditions over three consecutive
years of 1996-97, 1997-98 and 1998-99 are depicted in (Tables
1-3).
Morphological description of the somaclone "Parimal" and its wild
type parent
1) Genus -- Pelargonium 2) Species -- graveolens 3) Family --
Geraniaceae 4) Common name -- rose-scented geranium
TABLE-US-00002 TABLE 1 Comparative analysis of oil yield and
quality determining traits in the field grown morphologically
distinct somaclones, "Parimal" and wild type Pelargonium graveolens
CV. Hemanti (control). Soma- % contents of major terpenoids clone
Oil in the essential oil of geranium desig- Morpho- yield/ Cis +
trans nation types plant (g) Linalool rose oxide A Highly 4.5 .+-.
0.5 1.1 .+-. 0.2 1.6 .+-. 0.4 B dentated 3.3 .+-. 0.7 1.1 .+-. 0.2
1.3 .+-. 0.3 C leaf 8.6 .+-. 3.7 0.9 .+-. 0.1 1.2 .+-. 0.2 D 4.6
.+-. 1.2 1.0 .+-. 0.3 0.9 .+-. 0.3 Parimal Less 11.3 .+-. 2.8 9.1
.+-. 1.6 0.6 .+-. 0.12 dentated leaf L Less 8.0 .+-. 4.0 8.5 .+-.
2.8 0.6 .+-. 0.1 M dentated 4.5 .+-. 0.8 7.8 .+-. 1.5 0.7 .+-. 0.2
N leaf 5.1 .+-. 1.6 7.5 .+-. 1.3 0.6 .+-. 0.2 W Highly 3.8 .+-.
0.33 0.7 .+-. 0.1 1.5 .+-. 0.3 (control) dentated Soma- clone %
contents of major terpenoids desig- Morpho- in the essential oil of
geranium nation types Isomenthone Citronellol Geraniol A Highly 9.6
.+-. 1.8 49.8 .+-. 1.4 2.0 .+-. 0.5 B dentated 9.0 .+-. 2.5 52.6
.+-. 1.9 2.3 .+-. 0.6 C leaf 10.3 .+-. 1.3 51.7 .+-. 1.1 1.8 .+-.
0.4 D 10.6 .+-. 2.1 48.6 .+-. 4.5 2.3 .+-. 1.1 Parimal Less 7.8
.+-. 0.53 27.6 .+-. 2.1 22.6 .+-. 3.2 dentated leaf L Less 7.7 .+-.
0.5 28.7 .+-. 4.3 22.3 .+-. 2.8 M dentated 6.9 .+-. 1.5 29.2 .+-.
3.9 20.4 .+-. 2.1 N leaf 7.1 .+-. 1.2 29.1 .+-. 4.4 19.5 .+-. 3.1 W
Highly 9.5 .+-. 0.1 53.1 .+-. 0.7 1.6 .+-. 0.14 (control) dentated
% contents of major terpenoids Soma- in the essential oil of
geranium clone Citronel- desig- Morpho- llyl Geranyl 6,9-
10-epi-.quadrature.- nation types formate formate guaiadiene
eudesmol A Highly 13.0 .+-. 0.4 0.5 .+-. 0.3 1.3 .+-. 0.6 2.9 .+-.
0.5 B dentated 11.7 .+-. 1.3 0.3 .+-. 0.15 1.3 .+-. 0.4 3.0 .+-.
0.7 C leaf 11.1 .+-. 0.9 0.23 .+-. 0.03 1.3 .+-. 0.4 2.9 .+-. 0.5 D
10.2 .+-. 2.7 0.6 .+-. 0.45 1.2 .+-. 0.4 4.4 .+-. 1.4 Parimal Less
5.9 .+-. 0.2 2.1 .+-. 0.4 0.6 .+-. 0.12 7.3 .+-. 0.95 dentated leaf
L Less 5.6 .+-. 0.7 2.0 .+-. 0.2 0.7 .+-. 0.1 7.4 .+-. 1.1 M
dentated 4.8 .+-. 0.7 1.8 .+-. 0.2 0.7 .+-. 0.03 8.6 .+-. 1.8 N
leaf 4.7 .+-. 0.8 1.8 .+-. 0.6 0.6 .+-. 0.05 8.2 .+-. 1.6 W Highly
12.1 .+-. 0.6 0.23 .+-. 0.03 2.0 .+-. 0.2 3.2 .+-. 0.2 (control)
dentated
TABLE-US-00003 TABLE 2 Morphological characteristics of somaclone
"Parimal" and its wild type parent Morphological parameters Parimal
Wild type Plant height 85-95 cm 35-45 cm Growth habit spreading,
round canopy Spreading prostrate Stem green, internodes Greenish
magenta 8-9 cms long, purplish at the nodes, nodes pinkish 5-10 mm
in diameter, densely hairy Branches 91-101 10-45 Leaf Number per
branch 10-25 25-30 Texture pubescent pubescent Shape thick,
serrated with round thick hairy highly lobes serrated Number of
primary lobes 3 to 5 secondary lobes 3 15-30 12-15 Tip Blunt Blunt
Leaf area 20-28 cm.sup.2 30-36 cm.sup.2 Petiole .about.8 cm
(pinkish) .about.5 cm (green)
TABLE-US-00004 TABLE 3 Oil yield and quality determining traits of
somaclone "Parimal" and its wild type parent. Somaclone Trait
"Parimal" Wild type 1. Morphological marker Less dentated Highly
dentated leaf leaf morphotype morphotype (Hdl) (Ldl) 2. Leaf/stem
ratio (L/S) 2.1 1.5 3. Herb yield/plant 4.5 kgs. 1.9 kgs 4. % oil
content 0.18 0.15 5. Oil yield/plant 7.1 gms. 2.5 gms. 6. Oil yield
(kg/ha) 71.1 25.1 7. Oil quality i) Linalool 9.1 0.7 ii) Cis +
Trans rose 0.60 1.5 oxide iii) Menthone 0.2 0.3 iv) Isomenthone 7.8
9.5 v) Citronellol 27.6 53.1 vi) Geraniol 22.6 1.6 vii) Citronellyl
formate 5.9 12.1 viii) Geranyl formate 2.1 0.2 ix) Citronellyl
acetate 0.2 0.9 x) .beta.-caryophyllene 0.2 1.5 xi) 6,9 guaiadiene
0.6 2.0 xii) citronellyl butyrate 1.0 2.2 xiii) 10
Epi-.gamma.-eudesmol 7.3 3.2 xiv) Citronellyl tiglate 0.8 1.6
Type and frequency of variants during multiplication and
maintenance of the somaclone
No variants has been recovered during multiplication, maintenance
and testing of the somaclone "Parimal" in experimental plots,
screened carefully in 1996-97, 1997-98 and 1998-99 winter-summer
cropping seasons. The three year's observations on somaclone
"Parimal" were statistically analysed for significance of
differences in the morphological oil yield determining attributes
and the concentrations of 9 geranium oil quality determining
terpenoid constituents. The intraclonal variation in clone
"Parimal" was found statistically insignificant over three year's
field trial.
Statement of distinction
The somaclones "Parimal" are distinct in having large round canopy,
less dentated round leaf morphology, extensive branching and
absence of flowering (Table 1). Its oil profile is distinct from
the wild type parent cultivar in having higher proportion of
geraniol and lower proportion of citronellol, lower content of
cis+trans rose oxide and citronellyl formate, 6,9-guadiene, higher
content of linalool, geranyl formate and 10-epi-y endesmol
(Table-2). Its RAPD profile was different from the wild type parent
in certain aspects while had certain features common in both.
Characterization through RAPD markers
Using 13 random primers (MAP 01 to MAP 16) with following sequence
description a comparison was made for somaclone "Parimal" and its
wild type parent to establish the relatedness between them.
TABLE-US-00005 Primer Base Sequence MAP01 GTGCAATCAG SEQ. ID NO: 1
MAP02 AGGATACGTG SEQ. ID NO: 2 MAP04 AAGATAGCGG SEQ. ID NO: 3 MAP05
GGATCTGAAC SEQ. ID NO: 4 MAP06 TTGTCTCAGG SEQ. ID NO: 5 MAP07
GTCCTACTCG SEQ. ID NO: 6 MAP09 TGCGCGATCG SEQ. ID NO: 7 MAP10
AACGTACGCG SEQ. ID NO: 8 MAP11 GCACGCCGGA SEQ. ID NO: 9 MAP12
CACCCTGCGC SEQ. ID NO: 10 MAP14 GGACTCCACG SEQ. ID NO: 11 MAP15
AAGATAGCGG SEQ. ID NO: 12 MAP16 CTATCGCCGC SEQ. ID NO: 13
This analysis could narrow down with usage of six MAP primers (MAP
04, MAP 05, MAP 10, MAP 14, MAP 15, MAP 16) which in turn were
utilized for genotypic pair analysis in order to assess
genetic/molecular distances. The following table 4 gives the
similarity index between the two genotypes in paired test.
TABLE-US-00006 TABLE 4 Similarity percentage for amplified bands
(paired analysis) of somaclone "Parimal" and wild type parent
Primer Genotypic pair (somaclone "Parimal"/wild type) MAP 04 93 MAP
05 80 MAP 10 60 MAP 14 90 MAP 15 83 MAP 16 66
It is evident from similarity indices (paired test) that somaclone
"Parimal" and wild type parent are genetically distinguishable
(Table 4). Further, the primer MAP 10, MAP 14 showed least
similarity while primers MAP 04 and MAP15 showed close relatedness.
The primers MAP05 and MAP16 showed intermediate degree of
similarity. These observations on percent similarity confirm the
relatedness of somaclone "Parimal" with the wild type parent, at
the same time highlight the distinctness of the former at the
genetic level. The complete RAPD profile for PCR amplified DNA
segments using 12 primers is available providing evidence of
distinctiveness of the clone with respect to the control parent as
shown in FIG. 4.
Advantages:
1. "Parimal" is well adaptive to the unconventional area of
cultivation i.e. northern Indian plains. 2. "Parimal" represents
improvement over its wild type parental cv. "Hemanti" with resepect
to the following quantitative traits: (a) Larger and denser canopy;
(b) Greater herb yield; (c) Higher leaf:stem ratio; (d) Higher oil
content; and (e) Much higher oil yield (both on per plant and per
hectare basis). 3. "Parimal" represent improvement over its wild
type parental cv. "Hemanti" with respect to following major
essential oil quality determining traits: (a) About equal
citronellol:geraniol ratio; (b) Higher rhodinol content; (c) Higher
linalool concentration; (d) Lower concentration of 6,9-guaiadiene.
4. The somaclone "Parimal" showed consistency in performance for
all the above mentioned oil yield and quality determining
attributes during its field evaluation under Lucknow conditions
over three consecutive years. 5. "Parimal" showed distinctiveness
even at genetic level with respect to RAPD primers, MAP-05, MAP-12,
MAP-15 and MAP-16.
SEQUENCE LISTINGS
1
13110DNAartificial sequencerandom primers 1 gtgcaatcag 10
210DNAARTIFICIAL SEQUENCErandom primer 2 aggatacgtg 10
310DNAartificial sequencerandom primer 3 aagatagcgg 10
410DNAartificial sequencerandom primer 4 ggatctgaac 10
510DNAartificial sequencerandom primer 5 ttgtctcagg 10
610DNAartificial sequencerandom primer 6 gtcctactcg 10
710DNAartificial sequencerandom primer 7 tgcgcgatcg 10
810DNAartificial sequencerandom primer 8 aacgtacgcg 10
910DNAartificial sequencerandom primer 9 gcacgccgga 10
1010DNAartificial sequencerandom primer 10 caccctgcgc 10
1110DNAartificial sequencerandom primer 11 ggactccacg 10
1210DNAartificial sequencerandom primer 12 aagatagcgg 10
1310DNAartificial sequencerandom primer 13 ctatcgccgc 10
* * * * *