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Library:15076 (dbEST ID)
zmrsub1
Library Description
| Organism: | Zea mays |
(1) The zmrsub1 library was created by subtracting the well-watered maize root library zmrww00 from the water-starved maize root libraries zmrws05 and zmrws48. The 3 starting libraries are described below in (3). (2) Purified plasmid preps from zmrws05 and zmrws48 were combined. Sequenced clones from the zmrww00 library were pooled and converted into single-stranded circles and used as a driver for subtraction. The PCR products from the driver were hybridized with the single-stranded version of the zmrws05-zmrws48 library for 88 hours at 30 oC. Unhybridized single-stranded DNA circles were separated from hybridized DNA rendered partially double-stranded and electroporated into DH110B cells to generate the subtracted library. The total number of clones was 2 x 106 cfu. Background of empty clones (detected by blue/white selection on agar plates) was approximately 10%. Insert size of the subtracted library, determined by PCR of the entire library, ranges from 0.5 to 2.1kb). (3) Root samples were collected in Robert E. Sharp's lab (University of Missouri-Columbia) to construct three normalized cDNA libraries. Dark-grown maize seedlings with primary roots 12-20 mm in length were transplanted to high (-0.03 MPa) or low water potential (-1.6 MPa) vermiculite, and harvested at 5 h and 48 h after transplanting. About 1,000 roots were used for each of the low water potential libraries (zmrws05 and zmrws48) while 500 roots were combined from each of the two time points at high water potential (zmrww00). Each root was divided into 4 segments (distances are from the junction of the root apex and root cap): segment 1, 0-3 mm plus the root cap; segment 2, 3-7 mm; segment 3, 7-12 mm; segment 4, 12-20 mm. (For detatils of conditions see (1) with nutrient modifications as in (2)). The three normalized cDNA libraries were constructed in the lab of Hans Bohnert (University of Illinois-UC). Total RNA was extracted by the 'hot Phenol' method (Plant Molecular Biology manual. D5: 1-13, 2nd ed., 1997). This method worked in eliminating carbohydrate material present in the root tips. The integrity of the RNA was verified by denaturing agarose gels and spectrophotometry (ratio A260/280). Poly(A)+mRNA was isolated twice from total RNA using the Oligotex Direct mRNA kit (Qiagen). Poly(A)+ mRNA was converted to double-stranded cDNA and tagged by using modified Oligo(dT) primers. One of 4 sequence tags corresponding to a different segment of the root was added to the 3'-end of the modified Oligo(dT) primers, including a Not1 site and used to reverse transcribe the segment-specific mRNAs into cDNAs. Each library contains all four tags. A suffix (s1, s2, s3, or s4) has been added to each sequence identifier to designate which region of the root (Root_segment_1, 2, 3, or 4) the sequence was found in based on the identification of the tag. A suffix of s0 indicates that the sequence tag, and hence the source segment, could not be identified. The double stranded cDNAs were size-selected (>450 bp). Size selected cDNAs were adaptored with EcoRI adaptors at both ends, and then digested with NotI. The cDNA was directionally cloned into EcoR1-NotI digested pBS II SK(+) phagemid vector (Stratagene) and electroporated into E.coli DH10B. The total number of white colony forming units (cfu) in the primary libraries before amplification was as follows: zmrws05: 3.37 x 106; zmrws48: 4.87 x 106; zmrww00: 3 x 106. The background of empty clones was less than 1%. Inserts ranged from ~0.5kb to >2.5 kb, as determined by PCR. Plasmid DNA from the primary libraries then was converted to single-stranded circles and used as a template for PCR amplification using the T7 and T3 priming sites that flank the cloned cDNA inserts. The purified PCR products, representing the entire cDNA population cloned in each library, were used as a driver for normalization. Hybridization between the single-stranded library and the PCR products was carried out for 44 hours at 30oC. Non-hybridized single-stranded DNA circles were separated from hybridized DNA rendered partially double-stranded and electroporated into DH10B. The total number of clones with insert was: zmrws05: 2.0x107; zmrws48: 4.2x107; zmrww00: 1.1x107. The background of empty clones was less than 2%. Insert size, determined by PCR of the entire library, ranged from 0.5kb to 2.5kb. (1) Sharp R E; Silk W K; Hsiao T C. Growth of the Maize Primary Root at Low Water Potentials I. Spatial Distribution of Expansive Growth. Plant Physiology (Rockville). 87(1). 1988. 50-57. (2) Spollen W G; LeNoble M E; Samuels T D; Bernstein N; Sharp R E. Abscisic acid accumulation maintains maize primary root elongation at low water potentials by restricting ethylene production. Plant Physiology (Rockville). 122(3). March, 2000. 967-976.
Source: Hans Bohnert, University of Illinois, Urbana-Champaign
Gene Content Analysis
6,172 ESTs from this library were grouped into 4,149 UniGene entries (putative genes) [UniGene build #74, 02-Dec-2008]. EST counts for each entry may be used to calculate an approximate expression level in transcripts per million (TPM).
| ESTs | TPM | UniGene Entry | ||
|---|---|---|---|---|
| 28 | 4537 |  |
Zm.82832 | Clone 306068 unknown mRNA. |
| 21 | 3402 |  |
Zm.83125 | Clone 1471547 mRNA sequence. |
| 18 | 2916 | |
Zm.67921 | Clone 1470257 mRNA sequence. |
| 17 | 2754 | |
Zm.7000 | Clone 1468556 unknown mRNA. |
| 14 | 2268 |  |
Zm.8152 | Hypothetical protein LOC100192746 (LOC100192746), mRNA. |
| 13 | 2106 | |
Zm.84829 | Clone 303496 unknown mRNA. |
| 13 | 2106 | |
Zm.67193 | Clone 1684741 unknown mRNA. |
| 13 | 2106 | |
Zm.23253 | Histone H4 (H4C14), mRNA. |
| 13 | 2106 | |
Zm.1053 | Clone 1475432 mRNA sequence. |
| 12 | 1944 | |
Zm.74035 | Clone 1710772 mRNA sequence. |
| 11 | 1782 | |
Zm.98101 | Transcribed locus, weakly similar to NP_001047271.1 Os02g0587000 [Oryza sativa (japonica cultivar-group)]. |
| 11 | 1782 | |
Zm.93753 | Clone 451113 mRNA sequence. |
| 11 | 1782 | |
Zm.550 | Clone 1421744 unknown mRNA. |
| 10 | 1620 | |
Zm.9 | Clone 1587950 mRNA sequence. |
| 10 | 1620 | |
Zm.3634 | Clone 1349913 unknown mRNA. |
| 9 | 1458 |  |
Zm.93658 | Clone 1708726 unknown mRNA. |
| 9 | 1458 | |
Zm.68107 | Clone 1278371 unknown mRNA. |
| 9 | 1458 | |
Zm.12584 | Clone 1511555 mRNA sequence. |
| 8 | 1296 | |
Zm.82291 | Clone 450132 unknown mRNA. |
| 8 | 1296 | |
Zm.78127 | Clone 1478218 mRNA sequence. |
| 8 | 1296 | |
Zm.5678 | Clone 262650 unknown mRNA. |
| 8 | 1296 | |
Zm.2376 | Physical impedance induced protein (IIG1), mRNA. |
| 8 | 1296 | |
Zm.18097 | Transcribed locus, weakly similar to NP_001059904.1 Os07g0542900 [Oryza sativa (japonica cultivar-group)]. |
| 8 | 1296 | |
Zm.122075 | Clone 398088 mRNA sequence. |
| 7 | 1134 | |
Zm.92368 | Clone 1216993 unknown mRNA. |
| 7 | 1134 | |
Zm.9197 | Clone 1000184 unknown mRNA. |
| 7 | 1134 | |
Zm.87052 | Clone 1473944 mRNA sequence. |
| 7 | 1134 | |
Zm.82122 | Clone 1479543 mRNA sequence. |
| 7 | 1134 | |
Zm.5778 | Clone 1594526 unknown mRNA. |
| 7 | 1134 | |
Zm.355 | Clone 1468921 mRNA sequence. |
| 7 | 1134 | |
Zm.3347 | Clone 1529582 unknown mRNA. |
| 7 | 1134 | |
Zm.244 | Clone 1478051 unknown mRNA. |
| 7 | 1134 | |
Zm.19682 | Hypothetical protein LOC100193556 (LOC100193556), mRNA. |
| 7 | 1134 | |
Zm.169 | Clone 263279 unknown mRNA. |
| 6 | 972 | |
Zm.97076 | Hypothetical protein LOC100192585 (LOC100192585), mRNA. |
| 6 | 972 | |
Zm.96653 | Clone 1061239 unknown mRNA. |
| 6 | 972 | |
Zm.94015 | Clone 490573 mRNA sequence. |
| 6 | 972 | |
Zm.92749 | Clone 3535 mRNA sequence. |
| 6 | 972 | |
Zm.92695 | Clone 1515824 unknown mRNA. |
| 6 | 972 | |
Zm.84715 | Hypothetical protein LOC100191929 (LOC100191929), mRNA. |
| 6 | 972 | |
Zm.8130 | Clone 1379302 unknown mRNA. |
| 6 | 972 | |
Zm.81073 | Clone EL01N0438F11.c mRNA sequence. |
| 6 | 972 | |
Zm.80838 | Clone 490938 mRNA sequence. |
| 6 | 972 | |
Zm.80712 | Clone 275519 mRNA sequence. |
| 6 | 972 | |
Zm.79224 | Clone 1706016 mRNA sequence. |
| 6 | 972 | |
Zm.79043 | Clone 219489 unknown mRNA. |
| 6 | 972 | |
Zm.78368 | Lipoxygenase. |
| 6 | 972 | |
Zm.65545 | Transcribed locus, weakly similar to NP_001047046.1 Os02g0537100 [Oryza sativa (japonica cultivar-group)]. |
| 6 | 972 | |
Zm.64 | Clone 306156 mRNA sequence. |
| 6 | 972 | |
Zm.13947 | Hypothetical protein LOC100193883 (LOC100193883), mRNA. |
| 6 | 972 | |
Zm.119410 | Clone 1564857 unknown mRNA. |
| 6 | 972 | |
Zm.117489 | Clone 263905 unknown mRNA. |
| 6 | 972 | |
Zm.10825 | Clone 1468919 mRNA sequence. |
| 6 | 972 | |
Zm.103634 | Clone 1679916 unknown mRNA. |
| 5 | 810 |  |
Zm.97266 | Clone 326588 unknown mRNA. |
| 5 | 810 | |
Zm.96067 | Clone 1708356 mRNA sequence. |
| 5 | 810 | |
Zm.94897 | Clone 321354 unknown mRNA. |
| 5 | 810 | |
Zm.94847 | Clone 1468952 mRNA sequence. |
| 5 | 810 | |
Zm.94680 | Clone 1476811 mRNA sequence. |
| 5 | 810 | |
Zm.94353 | Hypothetical protein LOC100191627 (LOC100191627), mRNA. |
| 5 | 810 | |
Zm.94212 | Clone 1671141 mRNA sequence. |
| 5 | 810 | |
Zm.94101 | Clone 373282 unknown mRNA. |
| 5 | 810 | |
Zm.93949 | Clone 312744 unknown mRNA. |
| 5 | 810 | |
Zm.93929 | Clone 371243 unknown mRNA. |
| 5 | 810 | |
Zm.93833 | Transcribed locus, strongly similar to NP_001105440.1 Alpha-tubulin #3 [Zea mays]. |
| 5 | 810 | |
Zm.86906 | Hypothetical protein LOC100193874 (LOC100193874), mRNA. |
| 5 | 810 | |
Zm.82353 | Hypothetical protein LOC100191555 (LOC100191555), mRNA. |
| 5 | 810 | |
Zm.82164 | Clone 368653 mRNA sequence. |
| 5 | 810 | |
Zm.81281 | Hypothetical protein LOC100194265 (LOC100194265), mRNA. |
| 5 | 810 | |
Zm.79487 | Clone 219112 mRNA sequence. |
| 5 | 810 | |
Zm.74255 | Clone 278707 unknown mRNA. |
| 5 | 810 | |
Zm.70761 | Clone 208348 mRNA sequence. |
| 5 | 810 | |
Zm.68866 | Hypothetical protein LOC100193584 (LOC100193584), mRNA. |
| 5 | 810 | |
Zm.67270 | Clone 282963 mRNA sequence. |
| 5 | 810 | |
Zm.67234 | Clone 1291501 unknown mRNA. |
| 5 | 810 | |
Zm.67212 | Clone 1425326 unknown mRNA. |
| 5 | 810 | |
Zm.66639 | Clone 243478 unknown mRNA. |
| 5 | 810 | |
Zm.64864 | Clone 299390 unknown mRNA. |
| 5 | 810 | |
Zm.614 | Clone 687882 mRNA sequence. |
| 5 | 810 | |
Zm.6096 | Full-length cDNA clone ZM_BFb0073I17. |
| 5 | 810 | |
Zm.56887 | Clone 1471476 mRNA sequence. |
| 5 | 810 | |
Zm.549 | Clone 237341 unknown mRNA. |
| 5 | 810 | |
Zm.4932 | Clone 1477049 unknown mRNA. |
| 5 | 810 | |
Zm.4209 | Clone 221123 mRNA sequence. |
| 5 | 810 | |
Zm.4165 | Clone 1272474 unknown mRNA. |
| 5 | 810 | |
Zm.36063 | Clone 1613199 unknown mRNA. |
| 5 | 810 | |
Zm.29304 | Hypothetical protein LOC100192589 (LOC100192589), mRNA. |
| 5 | 810 | |
Zm.2193 | Clone 324412 mRNA sequence. |
| 5 | 810 | |
Zm.1894 | Clone 340475 mRNA sequence. |
| 5 | 810 | |
Zm.17964 | Clone 350091 unknown mRNA. |
| 5 | 810 | |
Zm.17336 | Transcribed locus, moderately similar to NP_001063525.1 Os09g0487500 [Oryza sativa (japonica cultivar-group)]. |
| 5 | 810 | |
Zm.16779 | Clone 232770 unknown mRNA. |
| 5 | 810 | |
Zm.122147 | Clone 238493 unknown mRNA. |
| 4 | 648 | |
Zm.99722 | Transcribed locus. |
| 4 | 648 | |
Zm.99677 | Transcribed locus, moderately similar to NP_001105100.1 benzoxazin6 [Zea mays]. |
| 4 | 648 | |
Zm.99669 | Clone 421771 unknown mRNA. |
| 4 | 648 | |
Zm.97923 | Transcribed locus, moderately similar to NP_001131436.1 hypothetical protein LOC100192768 [Zea mays]. |
| 4 | 648 | |
Zm.97434 | Full-length cDNA clone ZM_BFc0046F03. |
| 4 | 648 | |
Zm.96384 | Clone 352664 mRNA sequence. |
| 4 | 648 | |
Zm.95782 | Clone 396653 mRNA sequence. |
Source: NCBI UniGene
EST Sequences
6,458 EST sequences from this library have been submitted to the public sequence database as of 02-Dec-2008.
Source: NCBI GenBank
Click the button to save sequences to a file (FASTA format)
Keywords
- root
- seedling
- subtracted
Source: NCBI Clone Registry