Parent Washington Navel Orange Thrip-Challenged Flavedo cDNA Library UCRCS07

Overview
Library NameParent Washington Navel Orange Thrip-Challenged Flavedo cDNA Library UCRCS07
Unique NameParent Washington Navel Orange Thrip-Challenged Flavedo cDNA Library UCRCS07
OrganismCitrus sinensis (Sweet orange)
Typecdna_library
Vector: Lambda Uni-ZAP XR, excised phagemid; Site_1: EcoRI; Site_2: XhoI; Trees were grown in the field at University of California, Riverside using standard horticultural practices. Young fruits were placed in a cage with thrips (Scirtothrips citri). Infestations were conducted by Watkins (Morse lab) and flavedo collected by Federici (Roose lab) from May-June 2003. The thrips were collected from naturally infested Rhus plants by sucking into a tube. A flexible hose was attached to a tube that extended into a covered vial, and another tube stuck out of the vial at a right angle. The bent tube was held above the thrips, sucking on the flexible tube created a vacuum, pulling the thrips into the vial. Thrips were knocked off the Rhus plant onto a manila folder, then only second instars were captured. This was done repeatedly until enough were obtained. Approximately 7-10 thrips were caged on each fruit within a plastic vial made of a 8 cm long by 5 cm diameter tube that had a very fine mesh organdy fabric glued to the bottom. The plastic cap was slit from the edge to the center so it could be slipped over the stem of the fruit. It was put in place, the thrips were knocked into the vial and it was fastened onto the cap then all gaps were closed with masking tape. The thrips naturally move up to the fruit. The cages were left in place for two days, then removed. The fruit were checked to be sure the thrips had stayed on, and then brought to the lab to cut off the flavedo using a razor blade. Only the flavedo from the stem 1/3 to 1/2 of the fruit was used. For controls an equal number of comparable sized fruit were caged without thrips, and the peel collected from them in the same manner. Tissues were frozen in liquid nitrogen, then stored at -80C until further processing. Fenton (Close lab) purified RNA by the phenol method described in J. Japanese Soc. Hort. Sci. 1996. 64 (4): 809-814, purified poly(A) mRNA using an Oligotex mRNA Kit (Qiagen), produced a primary cDNA library using a lambda ZAP XR cDNA Synthesis Kit (Stratagene), then mass-excised 0.77 million pfu from the primary library to produce a phagemid population. The library was made from equal portions of RNA from each of the two treatments. Phagemids were plated, plasmid DNA purified, cDNA clones archived, and DNA sequences determined bi-directionally using an ABI3730 at DNA Landmarks (Landry, Hubert, Laforest, Landry, Ligonde). Chromatogram files were downloaded by FTP by Close, then processed by Wanamaker (Close lab) using the HarvEST pipeline (http://harvest.ucr.edu) to remove vector and cloning oligo sequences and various contaminants, and to trim to a high quality region. Sequences that retained a phred 17 region of at least 100 bases were assembled, then chimeras were removed following manual inspection of assemblies (Close, Roose, Federici, Wanamaker, Lyon, Ye, Jang, Collin, Kacar, Ikeda, Quinitio). Sequences that survived all removal steps were submitted to GenBank.
SNP Chip Base
Array NameParent Washington Navel Orange Thrip-Challenged Flavedo cDNA Library UCRCS07
OrganismCitrus sinensis (Sweet orange)
Typecdna_library
Features
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Feature NameUnique NameType
CX046721CX046721EST
CX046720CX046720EST
CX046719CX046719EST
CX046718CX046718EST
CX046717CX046717EST
CX046716CX046716EST
CX046715CX046715EST
CX046714CX046714EST
CX046713CX046713EST
CX046712CX046712EST
CX046711CX046711EST
CX046710CX046710EST
CX046709CX046709EST
CX046706CX046706EST
CX046705CX046705EST
CX046704CX046704EST
CX046703CX046703EST
CX046702CX046702EST
CX046701CX046701EST
CX046700CX046700EST
CX046699CX046699EST
CX046698CX046698EST
CX046697CX046697EST
CX046696CX046696EST
CX046695CX046695EST

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Properties
Property NameValue
Epicarptissue type
Genbank library cultivarParent Washington Navel
Genbank library dev stage11 year old trees
Genbank library noteVector: Lambda Uni-ZAP XR, excised phagemid; Site_1: EcoRI; Site_2: XhoI; Trees were grown in the field at University of California, Riverside using standard horticultural practices. Young fruits were placed in a cage with thrips (Scirtothrips citri). Infestations were conducted by Watkins (Morse lab) and flavedo collected by Federici (Roose lab) from May-June 2003. The thrips were collected from naturally infested Rhus plants by sucking into a tube. A flexible hose was attached to a tube that extended into a covered vial, and another tube stuck out of the vial at a right angle. The bent tube was held above the thrips, sucking on the flexible tube created a vacuum, pulling the thrips into the vial. Thrips were knocked off the Rhus plant onto a manila folder, then only second instars were captured. This was done repeatedly until enough were obtained. Approximately 7-10 thrips were caged on each fruit within a plastic vial made of a 8 cm long by 5 cm diameter tube that had a very fine mesh organdy fabric glued to the bottom. The plastic cap was slit from the edge to the center so it could be slipped over the stem of the fruit. It was put in place, the thrips were knocked into the vial and it was fastened onto the cap then all gaps were closed with masking tape. The thrips naturally move up to the fruit. The cages were left in place for two days, then removed. The fruit were checked to be sure the thrips had stayed on, and then brought to the lab to cut off the flavedo using a razor blade. Only the flavedo from the stem 1/3 to 1/2 of the fruit was used. For controls an equal number of comparable sized fruit were caged without thrips, and the peel collected from them in the same manner. Tissues were frozen in liquid nitrogen, then stored at -80C until further processing. Fenton (Close lab) purified RNA by the phenol method described in J. Japanese Soc. Hort. Sci. 1996. 64 (4): 809-814, purified poly(A) mRNA using an Oligotex mRNA Kit (Qiagen), produced a primary cDNA library using a lambda ZAP XR cDNA Synthesis Kit (Stratagene), then mass-excised 0.77 million pfu from the primary library to produce a phagemid population. The library was made from equal portions of RNA from each of the two treatments. Phagemids were plated, plasmid DNA purified, cDNA clones archived, and DNA sequences determined bi-directionally using an ABI3730 at DNA Landmarks (Landry, Hubert, Laforest, Landry, Ligonde). Chromatogram files were downloaded by FTP by Close, then processed by Wanamaker (Close lab) using the HarvEST pipeline (http://harvest.ucr.edu) to remove vector and cloning oligo sequences and various contaminants, and to trim to a high quality region. Sequences that retained a phred 17 region of at least 100 bases were assembled, then chimeras were removed following manual inspection of assemblies (Close, Roose, Federici, Wanamaker, Lyon, Ye, Jang, Collin, Kacar, Ikeda, Quinitio). Sequences that survived all removal steps were submitted to GenBank.
Genbank library tissue typeFlavedo