FRUIT QUALITY EVALUATION OF TRANSGENIC 'MEEKER' AND MAJOR CULTIVARS OF RED RASPBERRY GROWN IN THE PNW

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
212	1123	1160	100%

Progress 08/23/04 to 09/30/08

Outputs
Progress Report Objectives (from AD-416) Identify aroma compounds in 'Meeker' raspberry using gas chromatography/olfactometry-mass spectrometry technique. Quantitatively analyze the aroma and volatile composition of wild type 'Meeker' raspberry. Compare aroma and volatile composition of RBDV resistant transgenic raspberry lines with 'Meeker' wild type raspberry. Evaluate flavor quality by descriptive analysis and consumer preference and correlate sensory data with instrumental analysis. Correlate the volatile composition with metabolic pathways, and potentially relate the composition with raspberry genotypes. Approach (from AD-416) The most important flavor compounds will be identified using a gas chromatography/olfactometry-mass spectrometry technique, and odor-active compounds will be identified based on GC/MS, GC-IR, and literature reports. After identification, the compounds will be used as 'flavor mark' to evaluate RBDV resistant transgenic lines. The important aroma compounds will be quantified using dynamic headspace- GC-MS and solid phase micro-extraction GC-MS techniques. Sugar profile and organic acid profile will be measured by HPLC methods. Sensory evaluations will be conducted by expert panel and consumer preference tests, and data correlated with instrumental analysis. Flavor precursors and their conversion to aroma compounds will also be investigated using GC analysis, and structure examined with HPLC/MS/MS techniques. Anthocyanins will be analyzed using chromatography and antioxidant activity will be measured using oxygen radical absorbance capacity (ORAC). Documents SCA with OSU. Formerely 5358-22000-028-02S (8/2007). Significant Activities that Support Special Target Populations Raspberry bushy dwarf virus (RBDV) causes significant reduction in yield and crumbly fruit in raspberries, blackberries and raspberry-blackberry hybrids, and the only means of control is through development of resistant cultivars. Genetic modifications have been made to �Meeker� red raspberries in an effort to develop resistance to RBDV while maintaining the desirable characteristics of �Meeker�, including sweetness, aroma profile and machine harvestability. Fruit quality indicators, such as �Brix, titratable acidity, and sugar and organic acid profiles, were performed on fruit from wild-type and five transgenic lines of �Meeker� raspberries grown in Oregon and Washington for the years 2004 and 2005. Significant differences (p<0.05) were found between years for all tests performed. �Brix and titratable acidity were also significantly affected by locations, with berries grown in Washington having higher acid and lower �Brix than Oregon. Statistical analysis demonstrated that all five RBDV-resistant transgenic lines were similar to the wild type �Meeker� red raspberry. Many important raspberry aroma volatiles were quantified in the wild type and transgenic raspberry lines. Variations between some of the transgenic lines and wild type �Meeker� have been seen in some compounds; however, there is no evidence that they are statistically different. Experimental results showed that fruit quality is affected by season and growing environments. Additionally, the five transgenic lines behaved similarly to the wild-type �Meeker� raspberry. These results demonstrated that the transgenic RBDV-resistant lines of �Meeker� could serve as a replacement for the wild type �Meeker� red raspberry, provided that the fruit was accepted in the marketplace. Methods of ADODR monitoring included meetings, phone calls and e-mail.

Impacts
(N/A)

Publications

Progress 10/01/06 to 09/30/07

Outputs
Progress Report Objectives (from AD-416) Identify aroma compounds in 'Meeker' raspberry using gas chromatography/olfactometry-mass spectrometry technique. Quantitatively analyze the aroma and volatile composition of wild type 'Meeker' raspberry. Compare aroma and volatile composition of RBDV resistant transgenic raspberry lines with 'Meeker' wild type raspberry. Evaluate flavor quality by descriptive analysis and consumer preference and correlate sensory data with instrumental analysis. Correlate the volatile composition with metabolic pathways, and potentially relate the composition with raspberry genotypes. Approach (from AD-416) The most important flavor compounds will be identified using a gas chromatography/olfactometry-mass spectrometry technique, and odor-active compounds will be identified based on GC/MS, GC-IR, and literature reports. After identification, the compounds will be used as 'flavor mark' to evaluate RBDV resistant transgenic lines. The important aroma compounds will be quantified using dynamic headspace- GC-MS and solid phase micro-extraction GC-MS techniques. Sugar profile and organic acid profile will be measured by HPLC methods. Sensory evaluations will be conducted by expert panel and consumer preference tests, and data correlated with instrumental analysis. Flavor precursors and their conversion to aroma compounds will also be investigated using GC analysis, and structure examined with HPLC/MS/MS techniques. Anthocyanins will be analyzed using chromatography and antioxidant activity will be measured using oxygen radical absorbance capacity (ORAC). Documents SCA with OSU. Significant Activities that Support Special Target Populations This report serves to document research conducted under a specific cooperative agreement between ARS and Oregon State University. Additional details of research can be found in the report for the parent project 5358-22000-033-00D, Physiology, Biochemistry, and Genetic Improvement of Small Fruit Crops. Dr. Qian and collaborators conducted the following research towards the agreements objectives: Four raspberry cultivars (�Chilliwack�, �Tulameen�, �Willamette� and �Yellow Meeker�) grown in Washington during 2005 were compared to �Meeker� red raspberries, commercially grown in different Washington locations, for important flavor components, including: �brix, titratable acidity, sugar (fructose, glucose and sucrose) concentrations, organic acid (citric and malic acid) concentrations, volatile aroma compound concentration and chiral compound ratios. Sugar analysis (measured by HPLC) found sucrose levels to be low in �Meeker� raspberries, but these levels were considerably higher for the other cultivars studied. Thirty volatile aroma compounds were quantified using stir bar sorptive extraction (SBSE) paired with gas chromatography-mass spectrometry (GC-MS) . These compounds showed all of the cultivars to vary considerably from �Meeker�, particularly for �-ionone, (Z)-3-hexenol and linalool. Several chiral compounds were separated using a CyclosilB column to analyze the isomeric ratios of several aroma compounds. Variation from the �Meeker� cultivar for the chiral ratios was seen for 6-methyl-5-hepten-2-ol and linalool for all cultivars studied. Raspberry bushy dwarf virus (RBDV) causes significant reduction in yield and crumbly fruit in raspberries and raspberry-blackberry hybrids. There is no effective treatment once a plant is infected, and the only effective prevention is the use of resistant cultivars. Genetic modifications were made to �Meeker� red raspberries in order to impart resistance to RBDV while maintaining the desirable marketing characteristics of �Meeker�, including the sweetness, aroma profile and machine harvestability. These transgenic and wild-type �Meeker� plants were grown in Oregon and Washington and harvested in 2004 and 2005 for fruit analysis. Raspberries were analyzed for �brix, titratable acidity, volatile aroma compound concentration and chiral compound ratios. Year- to-year and site-to-site variations were seen for �brix and titratable acidity, with Oregon raspberries having slightly higher �brix and lower titratable acidity than Washington raspberries. None of the variations between the transgenic lines and wild-type �Meeker� seen within a site and year were repeated in the other site or year, indicating that the differences were due to environmental rather than genetic factors. Variations between harvest locations and season were larger than variation between wild-type �Meeker� and the transgenic lines. Chiral analysis revealed very little variation between lines, locations or years for the compounds studied. ADODR Statement: The ADODR met with the PI at meetings during the year and discussed results through phone calls and e-mail. Accomplishments Four raspberry cultivars (�Chilliwack�, �Tulameen�, �Willamette� and �Yellow Meeker�) grown in Washington during 2005 were compared to �Meeker� red raspberries, commercially grown in different Washington locations, for important flavor components, including: �brix, titratable acidity, sugar (fructose, glucose and sucrose) concentrations, organic acid (citric and malic acid) concentrations, volatile aroma compound concentration and chiral compound ratios. Sugar analysis (measured by HPLC) found sucrose levels to be low in �Meeker� raspberries, but these levels were considerably higher for the other cultivars studied. Thirty volatile aroma compounds ((Z)-3-hexenol, 4-isopropylbenzyl alcohol, 6- methyl-5-hepten-2-ol, 2-nonanol, hexanal, (E)-2-hexenal, (Z)-3-hexenyl acetate, ethyl hexanoate, methyl nonanoate, 2-heptanone, 2-nonanone, raspberry ketone, zingerone, �-octalactone, �-decalactone, para-cymene, geraniol, �-ionone, �-ionone, limonene, linalool, myrcene, nerol, �- phellandrene, �-pinene, sabinene, �-terpinene, �-terpinene, �-terpineol, terpinen-4-ol and terpinolene) were quantified using stir bar sorptive extraction (SBSE) paired with gas chromatography-mass spectrometry (GC-MS) . �-Ionone, �-ionone, geraniol, linalool, (Z)-3-hexenol and raspberry ketone are particularly useful in separating raspberry cultivars. These compounds showed all of the cultivars to vary considerably from �Meeker�, particularly for �-ionone, (Z)-3-hexenol and linalool. Several chiral compounds were separated using a CyclosilB column to analyze the isomeric ratios of several aroma compounds. Variation from the �Meeker� cultivar for the chiral ratios was seen for 6-methyl-5-hepten-2-ol and linalool for all cultivars studied. Raspberry bushy dwarf virus (RBDV) causes significant reduction in yield and crumbly fruit in raspberries and raspberry-blackberry hybrids. There is no effective treatment once a plant is infected, and the only effective prevention is the use of resistant cultivars. Genetic modifications were made to �Meeker� red raspberries in order to impart resistance to RBDV while maintaining the desirable marketing characteristics of �Meeker�, including the sweetness, aroma profile and machine harvestability. These transgenic and wild-type �Meeker� plants were grown in Oregon and Washington and harvested in 2004 and 2005 for fruit analysis. Raspberries were analyzed for �brix, titratable acidity, volatile aroma compound concentration and chiral compound ratios. Year- to-year and site-to-site variations were seen for �brix and titratable acidity, with Oregon raspberries having slightly higher �brix and lower titratable acidity than Washington raspberries. Thirty volatile aroma compounds ((Z)-3-hexenol, 4-isopropylbenzyl alcohol, 6-methyl-5-hepten-2- ol, 2-nonanol, hexanal, (E)-2-hexenal, (Z)-3-hexenyl acetate, ethyl hexanoate, methyl nonanoate, 2-heptanone, 2-nonanone, raspberry ketone, zingerone, �-octalactone, �-decalactone, para cymene, geraniol, �-ionone, �-ionone, limonene, linalool, myrcene, nerol, �-phellandrene, �-pinene, sabinene, �-terpinene, �-terpinene, �-terpineol, terpinen-4-ol and terpinolene) were quantified using stir bar sorptive extraction (SBSE) paired with gas chromatography-mass spectrometry (GC-MS). None of the variations between the transgenic lines and wild-type �Meeker� seen within a site and year were repeated in the other site or year, indicating that the differences were due to environmental rather than genetic factors. Variations between harvest locations and season were larger than variation between wild-type �Meeker� and the transgenic lines. Chiral analysis revealed very little variation between lines, locations or years for the compounds studied. Publications: 1. Flavor composition of Trangenic raspberry busgy dwarf virus resistant �Meeker� raspberries. Sara Malowicki, 2007. MS thesis for Oregon State University 2. FLAVOR COMPOSITION OF RASPBERRY CULTIVARS GROWN IN THE PACIFIC NORTHWEST, 2007, to be Submitted to Journal of Agricultural and Food Chemistry, Sarah M.M. Malowicki, Michael C. Qian, and Robert Martin 3. FLAVOR QUALITY OF RASPBERRY BUSHY DWARF VIRUS-RESISTANT TRANSGENIC �MEEKER� RED RASPBERRIES, 2007, to be Submitted to J. Agric. Food Chem. Sarah M.M. Malowicki, Michael C. Qian, Robert Martin

Impacts
(N/A)

Publications

Progress 10/01/05 to 09/30/06

Outputs
Progress Report 4d Progress report. This report serves to document research conducted under a specific cooperative agreement between ARS and Oregon State University. Additional details of research can be found in the report for the parent project 5358-22000-028-00D, Integrated Management of Virus Diseases of Small Fruit Crops. Dr. Michael Qian and collaborators conducted the following research towards the agreements objectives: Raspberry bushy dwarf virus (RBDV) causes significant reduction in yield and crumbly fruit in raspberries, blackberries and raspberry-blackberry hybrids, and the only means of control is through development of resistant cultivars. Genetic modifications have been made to Meeker red raspberries in an effort to develop resistance to RBDV while maintaining the desirable characteristics of Meeker including sweetness, aroma profile and machine harvestability. Fruit quality indicators such as Brix, titratable acidity, and sugar and organic acid profiles were performed on fruit from wild-type and five transgenic lines of Meeker raspberries grown in Oregon and Washington for the years 2004 and 2005. Significant differences (p<0.05) were found between years for all tests performed. Brix and titratable acidity were also significantly affected by locations with berries grown in Washington having higher acid and lower Brix than Oregon. Statistical analysis demonstrated that all five RBDV-resistant transgenic lines were similar to the wild type 'Meeker' red raspberry. Many important raspberry aroma volatiles were quantified in the wild type and transgenic raspberry lines and variations between some of the transgenic lines and wild type Meeker have been seen in some compounds; however, there is no evidence that they are statistically different. Experimental results showed that fruit quality is affected by season and growing environments. Additionally, the five transgenic lines behaved similarly to the wild-type Meeker raspberry. These results demonstrated that the transgenic RBDV-resistant lines of Meeker could serve as a replacement for the wild type Meeker red raspberry provided that the fruit was accepted in the marketplace.

Impacts
(N/A)

Publications

Progress 10/01/04 to 09/30/05

Outputs
4d Progress report. This report serves to document research conducted under a specific cooperative agreement between ARS and Oregon State University. Additional details of research can be found in the report for the parent project 5358-22000-028-00D Integrated Management of Virus Diseases of Small Fruit Crops. Specific Objectives: The objectives of this study were to identify aroma compounds in 'Meeker' raspberry and compare aroma and volatile composition of RBDV resistant transgenic raspberry lines with 'Meeker' wild type raspberry. Results : Seventy-five aromas were identified. The potentially important aroma compounds in wild type 'Meeker' raspberry included 2,5-dimethyl-4-hydroxy- 3-(2H)-furanone, hexanal, alpha-ionone, beta-ionone, beta-damascenone, cis-3-hexenal, methional, cis-3-hexenol, linalool, butanoic acid, ethyl 2- methylpropanoate, geraniol, and 4-(p-hydroxyphenyl)-2-butanone. The finding was published in J. Agric. Food Chem. 2004, 52, 5155-5161). Comparison of volatile composition of transgenic lines with wild type 'Meeker' raspberry from 2003 season has been attempted. Seventy-six compounds were identified by GC-MS. Preliminary results showed that out of the five transgenic lines, some variations were observed in some lines. It appeared that there were three transgenic lines having similar volatile compositions as the wild types. The results were presented at IFT and ACS meetings and a conference proceedings article was submitted. Fruits from 2004 and 2005 are under study to confirm the results. The sugar and organic acid profiles were studied for 2004 fruits. The raspberries showed similar sugar concentrations. The concentrations of fructose, glucose and sucrose in all transgenic lines were similar to that of wild type 'Meeker' although there is a trend for Washington berries to have slightly higher fructose and sucrose concentrations while the Oregon varieties have more sucrose. Citric acid was the primary acid found in raspberries, but there was a significant amount of malic acid in all varieties. Citric acid concentrations varied between 1.24% and 1.75% with an average of 1.52%. Malic acid ranged from 0.18% to 0.36% with an average of 0.24%. More wild type 'Meeker' will be analyzed to verify that there is any statistically significant difference among these cultivars.

Impacts
(N/A)

Publications