Gene 315
BLASTx Output


BLASTx Output. The query sequence is the Deltagen target gene.  The top "hits" are shown with
accession number, BLAST score and probability.  The alignment of the best hit is also shown.

BLAST DATE : 3/13/2003

 

                                                                         Score    E
 
 
gi|6755813|ref|NP_036035.1|  toll-like receptor 2 [Mus muscu...  1447   0.0  
gi|21955346|gb|AAH14693.1|  similar to toll-like receptor 2 ...  1445   0.0  
gi|12836693|dbj|BAB23770.1|  data source:MGDsource key:MGI...  1443   0.0  
gi|19718734|ref|NP_003255.2|  toll-like receptor 2; toll/int...  1050   0.0  
 
>gi|6755813|ref|NP_036035.1| toll-like receptor 2 [Mus musculus]
 gi|20140895|sp|Q9QUN7|TLR2_MOUSE Toll-like receptor 2 precursor
 gi|5669820|gb|AAD46481.1|AF124741_1 Toll-like receptor 2 [Mus musculus]
 gi|5732910|gb|AAD49335.1|AF165189_1 toll-like receptor 2 [Mus musculus]
 gi|6120052|gb|AAF04277.1|AF185284_1 Toll-like receptor 2 [Mus musculus]
 gi|6760423|gb|AAF28345.1|AF216289_1 Toll-like receptor 2 [Mus musculus]
          Length = 784
 
 Score = 1447 bits (3746)Expect = 0.0
 
 Identities = 728/784 (92%)Positives = 728/784 (92%)
 Frame = +1
 
Query: 316  MLRALWLFWILVAITVLFSKRCSAQESLSCDASGVCDGRSRSFTSIPSGLTAAMKSLDLS 495
            MLRALWLFWILVAITVLFSKRCSAQESLSCDASGVCDGRSRSFTSIPSGLTAAMKSLDLS
Sbjct: 1    MLRALWLFWILVAITVLFSKRCSAQESLSCDASGVCDGRSRSFTSIPSGLTAAMKSLDLS 60
 
Query: 496  FNKITYIGHGDLRACANLQVLILKSSRINTIEGDAFYSLGXXXXXXXXXXXXXXXXXXXX 675
            FNKITYIGHGDLRACANLQVLILKSSRINTIEGDAFYSLG                    
Sbjct: 61   FNKITYIGHGDLRACANLQVLILKSSRINTIEGDAFYSLGSLEHLDLSDNHLSSLSSSWF 120
 
Query: 676  XXXXXXKYLNLMGNPYQTLGVTSLFPNLTNLQTLRIGNVETFSEIRRIDFAGLTSLNELE 855
                  KYLNLMGNPYQTLGVTSLFPNLTNLQTLRIGNVETFSEIRRIDFAGLTSLNELE
Sbjct: 121  GPLSSLKYLNLMGNPYQTLGVTSLFPNLTNLQTLRIGNVETFSEIRRIDFAGLTSLNELE 180
 
Query: 856  IKALSLRNYQSQSLKSIRDIHHLTLHLSESAFLLEIFADILSSVRYLELRDTNLARFQFS 1035
            IKALSLRNYQSQSLKSIRDIHHLTLHLSESAFLLEIFADILSSVRYLELRDTNLARFQFS
Sbjct: 181  IKALSLRNYQSQSLKSIRDIHHLTLHLSESAFLLEIFADILSSVRYLELRDTNLARFQFS 240
 
Query: 1036 PLPVDEVSSPMKKLAFRGSVLTDESFNXXXXXXXXXXXXXXVEFDDCTLNGLGDFNPSES 1215
            PLPVDEVSSPMKKLAFRGSVLTDESFN              VEFDDCTLNGLGDFNPSES
Sbjct: 241  PLPVDEVSSPMKKLAFRGSVLTDESFNELLKLLRYILELSEVEFDDCTLNGLGDFNPSES 300
 
Query: 1216 DVVSELGKVETVTIRRLHIPQFYLFYDLSTVYSLLEKVKRITVENSKVFLVPCSFSQHLK 1395
            DVVSELGKVETVTIRRLHIPQFYLFYDLSTVYSLLEKVKRITVENSKVFLVPCSFSQHLK
Sbjct: 301  DVVSELGKVETVTIRRLHIPQFYLFYDLSTVYSLLEKVKRITVENSKVFLVPCSFSQHLK 360
 
Query: 1396 SLEFLDLSENLMVEEYLKNSACKGAWPSLQTLVLSQNHLRSMQKTGEILLTLKNLTSLDI 1575
            SLEFLDLSENLMVEEYLKNSACKGAWPSLQTLVLSQNHLRSMQKTGEILLTLKNLTSLDI
Sbjct: 361  SLEFLDLSENLMVEEYLKNSACKGAWPSLQTLVLSQNHLRSMQKTGEILLTLKNLTSLDI 420
 
Query: 1576 SRNTFHPMPDSCQWPEKMRFLNLSSTGIRVVKTCIPQTLEVLDVSNNNLDSFSLFLPRLQ 1755
            SRNTFHPMPDSCQWPEKMRFLNLSSTGIRVVKTCIPQTLEVLDVSNNNLDSFSLFLPRLQ
Sbjct: 421  SRNTFHPMPDSCQWPEKMRFLNLSSTGIRVVKTCIPQTLEVLDVSNNNLDSFSLFLPRLQ 480
 
Query: 1756 ELYISRNKLKTLPDASLFPVLLVMKIRENAVSTFSKDQLGSFPKLETLEAGDNHFVCSCE 1935
            ELYISRNKLKTLPDASLFPVLLVMKIRENAVSTFSKDQLGSFPKLETLEAGDNHFVCSCE
Sbjct: 481  ELYISRNKLKTLPDASLFPVLLVMKIRENAVSTFSKDQLGSFPKLETLEAGDNHFVCSCE 540
 
Query: 1936 LLSFTMETPALAQILVDWPDSYLCDSPPRLHGHRLQDARPSVLECHQAALVSXXXXXXXX 2115
            LLSFTMETPALAQILVDWPDSYLCDSPPRLHGHRLQDARPSVLECHQAALVS        
Sbjct: 541  LLSFTMETPALAQILVDWPDSYLCDSPPRLHGHRLQDARPSVLECHQAALVSGVCCALLL 600
 
Query: 2116 XXXXXXXXCHHFHGLWYLRMMWAWLQAKRKPKKAPCRDVCYDAFVSYSEQDSHWVENLMV 2295
                    CHHFHGLWYLRMMWAWLQAKRKPKKAPCRDVCYDAFVSYSEQDSHWVENLMV
Sbjct: 601  LILLVGALCHHFHGLWYLRMMWAWLQAKRKPKKAPCRDVCYDAFVSYSEQDSHWVENLMV 660
 
Query: 2296 QQLENSDPPFKLCLHKRDFVPGKWIIDNIIDSIEKSHKTVFVLSENFVRSEWCKYELDFS 2475
            QQLENSDPPFKLCLHKRDFVPGKWIIDNIIDSIEKSHKTVFVLSENFVRSEWCKYELDFS
Sbjct: 661  QQLENSDPPFKLCLHKRDFVPGKWIIDNIIDSIEKSHKTVFVLSENFVRSEWCKYELDFS 720
 
Query: 2476 HFRLFDENNDAAILVLLEPIERKAIPQRFCKLRKIMNTKTYLEWPLDEGQQEVFWVNLRT 2655
            HFRLFDENNDAAILVLLEPIERKAIPQRFCKLRKIMNTKTYLEWPLDEGQQEVFWVNLRT
Sbjct: 721  HFRLFDENNDAAILVLLEPIERKAIPQRFCKLRKIMNTKTYLEWPLDEGQQEVFWVNLRT 780
 
Query: 2656 AIKS 2667
            AIKS
Sbjct: 781  AIKS 784
 
 
 

Construct

Gene:

315

 

GI Number(s):

6760422

ES Cell Line:

170

 

 

Accession Number:

AF216289

 

 

Family:

Interleukin/Toll Receptor

 

 

Subfamily:

Toll-like

 

 

Species:

Mouse

 

 

 

Sequence Type:

full length cDNA

 

 

 

 

 


Sequence Length:

2838 bp

 

red underlined text = deleted in gene sequence

Sequence Deleted:

from base 1678 to base 2094

 

black text = gene sequence flanking deleted sequence



GGCACGAGAGATGGCTAGTGGGCACGGGGAGCGGCGGCTGGAGGACTCCTAGGCTCCGGG
CAGGCGGTCACTGGCAGGAGATGTGTCCGCAATCATAGTTTCTGATGGTGAAGGTTGGAC
GGCAGTCTCTGCGACCTAGAAGTGGAAAAGATGTCGTTCAAGGAGGTGCGGACTGTTTCC
TTCTGACCAGGATCTTGTTTCTGAGTGTAGGGGCTTCACTTCTCTGCTTTTCGTTCATCT
CTGGAGCATCCGAATTGCATCACCGGTCAGAAAACAACTTACCGAAACCTCAGACAAAGC
GTCAAATCTCAGAGGATGCTACGAGCTCTTTGGCTCTTCTGGATCTTGGTGGCCATAACA
GTCCTCTTCAGCAAACGCTGTTCTGCTCAGGAGTCTCTGTCATGTGATGCTTCTGGGGTG
TGTGATGGCCGCTCCAGGTCTTTCACCTCTATTCCCTCCGGACTCACAGCAGCCATGAAA
AGCCTTGACCTGTCTTTCAACAAGATCACCTACATTGGCCATGGTGACCTCCGAGCGTGT
GCGAACCTCCAGGTTCTGATTTTGAAGTCCAGCAGAATCAATACAATAGAGGGAGACGCC
TTTTATTCTCTGGGCAGTCTTGAACATTTGGATTTGTCTGATAATCACCTATCTAGTTTA
TCTTCCTCCTGGTTCGGGCCCCTTTCCTCTTTGAAATACTTAAACTTAATGGGAAATCCT
TACCAGACACTGGGGGTAACATCGCTTTTTCCCAATCTCACAAATTTACAAACCCTCAGG
ATAGGAAATGTAGAGACTTTCAGTGAGATAAGGAGAATAGATTTTGCTGGGCTGACTTCT
CTCAATGAACTTGAAATTAAGGCATTAAGTCTCCGGAATTATCAGTCCCAAAGTCTAAAG
TCGATCCGCGACATCCATCACCTGACTCTTCACTTAAGCGAGTCTGCTTTCCTGCTGGAG
ATTTTTGCAGATATTCTGAGTTCTGTGAGATATTTAGAACTAAGAGATACTAACTTGGCC
AGGTTCCAGTTTTCACCACTGCCCGTAGATGAAGTCAGCTCACCGATGAAGAAGCTGGCA
TTCCGAGGCTCGGTTCTCACTGATGAAAGCTTTAACGAGCTCCTGAAGCTGTTGCGTTAC
ATCTTGGAACTGTCGGAGGTAGAGTTCGACGACTGTACCCTCAATGGGCTCGGCGATTTC
AACCCCTCGGAGTCAGACGTAGTGAGCGAGCTGGGTAAAGTAGAAACAGTCACTATCCGG
AGGTTGCATATCCCCCAGTTCTATTTGTTTTATGACCTGAGTACTGTCTATTCCCTCCTG
GAGAAGGTGAAGCGAATCACAGTAGAGAACAGCAAGGTCTTCCTGGTTCCCTGCTCGTTC
TCCCAGCATTTAAAATCATTAGAATTCTTAGACCTCAGCGAAAATCTGATGGTTGAAGAA
TATTTGAAGAACTCAGCCTGTAAGGGAGCCTGGCCTTCTCTACAAACCTTAGTTTTGAGC
CAGAATCATTTGAGATCAATGCAAAAAACAGGAGAGATTTTGCTGACTCTGAAAAACCTG
ACCTCCCTTGACATCAGCAGGAACACTTTTCATCCGATGCCCGACAGCTGTCAGTGGCCA
GAAAAGATGCGCTTCCTGAATTTGTCCAGTACAGGGATCCGGGTGGTAAAAACGTGC
ATT
CCTCAGACGCTGGAGGTGTTGGATGTTAGTAACAACAATCTTGACTCATTTTCTTTGTTC
TTGCCTCGGCTGCAAGAGCTCTATATTTCCAGAAATAAGCTGAAAACACTCCCAGATGCT
TCGTTGTTCCCTGTGTTGCTGGTCATGAAAATCAGAGAGAATGCAGTAAGTACTTTCTCT
AAAGACCAACTTGGTTCTTTTCCCAAACTGGAGACTCTGGAAGCAGGCGACAACCACTTT
GTTTGCTCCTGCGAACTCCTATCCTTTACTATGGAGACGCCAGCTCTGGCTCAAATCCTG
GTTGACTGGCCAGACAGCTACCTGTGTGACTCTCCGCCTCGCCTGCACGGCCACAGGCTT
CAGGATGCCCGGCCCTCCGTCTTGGAATGTCACCAGGCTGCACTGGTGTCTGGA
GTCTGC
TGTGCCCTTCTCCTGTTGATCTTGCTCGTAGGTGCCCTGTGCCACCATTTCCACGGACTG
TGGTACCTGAGAATGATGTGGGCGTGGCTCCAGGCCAAGAGGAAGCCCAAGAAAGCTCCC
TGCAGGGACGTTTGCTATGATGCCTTTGTTTCCTACAGTGAGCAGGATTCCCATTGGGTG
GAGAACCTCATGGTCCAGCAGCTGGAGAACTCTGACCCGCCCTTTAAGCTGTGTCTCCAC
AAGCGGGACTTCGTTCCGGGCAAATGGATCATTGACAACATCATCGATTCCATCGAAAAG
AGCCACAAAACTGTGTTCGTGCTTTCTGAGAACTTCGTACGGAGCGAGTGGTGCAAGTAC
GAACTGGACTTCTCCCACTTCAGGCTCTTTGACGAGAACAACGACGCGGCCATCCTTGTT
TTGCTGGAGCCCATTGAGAGGAAAGCCATTCCCCAGCGCTTCTGCAAACTGCGCAAGATA
ATGAACACCAAGACCTACCTGGAGTGGCCCTTGGATGAAGGCCAGCAGGAAGTGTTTTGG
GTAAATCTGAGAACTGCAATAAAGTCCTAGGTTCTCCACCCAGTTCCTGACTTCCTTAAC
TAAGGTCTTTGTGACACAAACTGTAACAAAGTTTATAAGTAACATAGAATTGTATTATTG
AGGATATTAACTATGGGTTTTGTCTTGAATACTGTTATATAAATATGTGACATCAGGAAA
AAAAAAAAAAAAAAAAAA




Targeting Vector* (genomic sequence)

    Neo

Construct Number:  198

Arm Length:
5':  2.5  kb
3':  4  kb


  * Not drawn to scale



 

5 ' PCR Reconfirmation

Gene:

315

ES Line:

170

Gene Specific Oligo (GS):
ATGCTTTGCATCTATGAGCAGAGCA

12190



Gel

Expected Results and Diagram

 11000bp

 10000bp

 9000bp

 8000bp

 7000bp

 6000bp

 5000bp

 4000bp

 3000bp

 2000bp

 1600bp

 1000bp

 500bp

Lane Number

Label

Expected Size (in bp)

1

1kB Ladder

 

2

12190 + NEO1 (841)

3825

3

12190 + NEO2 (1446)

3200

4

12190 + NEO3 (1165)

2600

5

DNA Sample Control

5167

6

Positive Control

2500

 


 

3 ' PCR Reconfirmation

Gene:

315

ES Line:

170

Gene Specific Oligo (GS):
TTCTGGCCCTGGGGTCACAGAAGGG

1889



Gel

Expected Results and Diagram

 11000bp

 10000bp

 9000bp

 8000bp

 7000bp

 6000bp

 5000bp

 4000bp

 3000bp

 2000bp

 1600bp

 1000bp

 500bp

Lane Number

Label

Expected Size (in bp)

1

1kB Ladder

 

2

1889 + NEO1 (1431)

5000

3

1889 + NEO2 (1432)

4950

4

1889 + NEO3 (1170)

4250

5

DNA Sample Control

5167

6

Positive Control

4000

 


 

Legend : Genomic DNA from the recombinant ES line was assayed for homologous recombination using polymerase chain reactions (PCRs). Amplified DNA fragments were visualized by ethidium bromide staining following agarose gel electrophoresis, as shown in the gel image on the left. The test PCRs (a set of three lanes) employed the gene-specific (GS) primer, which lies outside of and adjacent to the targeting vector arm, paired in succession with one of three primers in the insertion fragment. The "DNA sample control" employed a primer pair intended to amplify a fragment from a non-targeted genomic locus. The "positive control" employed the GS primer paired with a primer at the other end of the arm. The table below the gel image lists the primers (numbered) used in each PCR and the expected product sizes, in base pairs (bp).  The diagrams to the right depict the anticipated gel image as well as the relative positions of the PCR primers.

 

 

 

5 ' Southern Blot Confirmation

Gene:

315

ES Line:

170

Construct:

198

Blot Number:

2147

Gene Side:

5'

 

 

MW Marker

Lane

ES Line

Enzyme

 

1

Parent 1

Eco RV

 

2

Parent 2

Eco RV

 

3

170

Eco RV

 

4

Additional 1

Eco RV

 

Legend : Genomic DNA isolated from the indicated ES lines was digested with the indicated restriction enzymes (determined to cut outside of the construct arms).  The DNA was analyzed by Southern hybridization, probing with a radiolabeled DNA fragment that hybridizes outside of and adjacent to the construct arm.  The parent ES lines (negative controls) showed bands representing the endogenous (wild-type) Gene #315 allele.  In contrast, the ES#170  line showed an additional band representing the targeted allele from the expected homologous recombination event.   "Additional" denotes a secondary line that was derived from the same targeting vector but was not used to generate mice.
Note: All lanes represent data from the same blot.

 

Genotyping: Gene 315 (F2 Mice)



 

W = wild-type

H = heterozygote

K = homozygote



A) Scheme:

B) Expected Result:

 

C) Oligo Information:

Type

ID

Sequence

Multiplex Conc.

GS(E,T)

3518

CTTCCTGAATTTGTCCAGTACAGGG

200

NEO(T)

3195

GGGCCAGCTCATTCCTCCCACTCAT

200

GS(E1)

3519

TCGACCTCGATCAACAGGAGAAGGG

400

 

D) Reaction Conditions:

Allele

Expected Size

Oligos to use

Targeted

320

oligo 3195 + oligo 3518

Endogenous

460

oligo 3519 + oligo 3518