Gene: 565 Name: Capn7 Family: Protease Subfamily: Cysteine Accession: NM_009796 GI: 6753257
Gene: 565

Gene 565
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|6753258|ref|NP_033926.1|  calpain 7 [Mus musculus] >gi|51...  1672   0.0  
gi|7656959|ref|NP_055111.1|  calpain 7; calpain like proteas...  1603   0.0  
gi|4495099|emb|CAB39203.1|  Capn7 [Mus musculus]                 1588   0.0  
gi|6680846|ref|NP_031628.1|  calpain 5 [Mus musculus] >gi|20...   124   1e-27
 
>gi|6753258|ref|NP_033926.1| calpain 7 [Mus musculus]
 gi|5102946|dbj|BAA78731.1| PalBH [Mus musculus]
          Length = 813
 
 Score = 1672 bits (4329)Expect = 0.0
 
 Identities = 813/813 (100%)Positives = 813/813 (100%)
 Frame = +3
 
Query: 159  MDASALERDAVQFARLAVQRDHEGRYSEAVFYYKEAAQALIYAEMAGSSLERIQEKINEY 338
            MDASALERDAVQFARLAVQRDHEGRYSEAVFYYKEAAQALIYAEMAGSSLERIQEKINEY
Sbjct: 1    MDASALERDAVQFARLAVQRDHEGRYSEAVFYYKEAAQALIYAEMAGSSLERIQEKINEY 60
 
Query: 339  LERVQALHSAVQSKSTDPLKSKHQLDLERAHFLVTQAFDEDEKGNVEDAIELYTEAVELC 518
            LERVQALHSAVQSKSTDPLKSKHQLDLERAHFLVTQAFDEDEKGNVEDAIELYTEAVELC
Sbjct: 61   LERVQALHSAVQSKSTDPLKSKHQLDLERAHFLVTQAFDEDEKGNVEDAIELYTEAVELC 120
 
Query: 519  LKTSSETADKTLQNKLKQLARQALDRAEALSEPLTKPFCKLKSANMKTKTPPVRTHFPLG 698
            LKTSSETADKTLQNKLKQLARQALDRAEALSEPLTKPFCKLKSANMKTKTPPVRTHFPLG
Sbjct: 121  LKTSSETADKTLQNKLKQLARQALDRAEALSEPLTKPFCKLKSANMKTKTPPVRTHFPLG 180
 
Query: 699  PNPFVEKPQAFISPQSCDAQGQKYTAEEIEVLRTTSKINGVEYVPFMSVDLRERFAYPMP 878
            PNPFVEKPQAFISPQSCDAQGQKYTAEEIEVLRTTSKINGVEYVPFMSVDLRERFAYPMP
Sbjct: 181  PNPFVEKPQAFISPQSCDAQGQKYTAEEIEVLRTTSKINGVEYVPFMSVDLRERFAYPMP 240
 
Query: 879  FCDRLGKLPLSPKQKTTFSKWVRPEDLTNNPTMIYTVSSFSIKQTIVSDCSFVASLAISA 1058
            FCDRLGKLPLSPKQKTTFSKWVRPEDLTNNPTMIYTVSSFSIKQTIVSDCSFVASLAISA
Sbjct: 241  FCDRLGKLPLSPKQKTTFSKWVRPEDLTNNPTMIYTVSSFSIKQTIVSDCSFVASLAISA 300
 
Query: 1059 AYERRFNKKLITSIIYPQNKDGEPEYNPCGKYMVKLHLNGVPRKVIIDDQLPVDHKGELL 1238
            AYERRFNKKLITSIIYPQNKDGEPEYNPCGKYMVKLHLNGVPRKVIIDDQLPVDHKGELL
Sbjct: 301  AYERRFNKKLITSIIYPQNKDGEPEYNPCGKYMVKLHLNGVPRKVIIDDQLPVDHKGELL 360
 
Query: 1239 CSYSNNKSELWVSLIEKAYMKVMGGYDFPGSNSNIDLHALTGWIPERIAMHSDSQTFSKD 1418
            CSYSNNKSELWVSLIEKAYMKVMGGYDFPGSNSNIDLHALTGWIPERIAMHSDSQTFSKD
Sbjct: 361  CSYSNNKSELWVSLIEKAYMKVMGGYDFPGSNSNIDLHALTGWIPERIAMHSDSQTFSKD 420
 
Query: 1419 NSFRMLYQRFHKGDVLITASTGVMTEAEGEKWGLVPTHAYAVLDIREFKGLRFIQLKNPW 1598
            NSFRMLYQRFHKGDVLITASTGVMTEAEGEKWGLVPTHAYAVLDIREFKGLRFIQLKNPW
Sbjct: 421  NSFRMLYQRFHKGDVLITASTGVMTEAEGEKWGLVPTHAYAVLDIREFKGLRFIQLKNPW 480
 
Query: 1599 SHLRWKGRYSENDVKNWTPELQKYLNFDPRTAQKIDNGIFWISWDDLCQYYDVVYLSWNP 1778
            SHLRWKGRYSENDVKNWTPELQKYLNFDPRTAQKIDNGIFWISWDDLCQYYDVVYLSWNP
Sbjct: 481  SHLRWKGRYSENDVKNWTPELQKYLNFDPRTAQKIDNGIFWISWDDLCQYYDVVYLSWNP 540
 
Query: 1779 ALFKESTCIHSTWDAKQGPVKDAYSLANNPQYKLEVQCPQGGAAVWVLLSRHITDKDDFA 1958
            ALFKESTCIHSTWDAKQGPVKDAYSLANNPQYKLEVQCPQGGAAVWVLLSRHITDKDDFA
Sbjct: 541  ALFKESTCIHSTWDAKQGPVKDAYSLANNPQYKLEVQCPQGGAAVWVLLSRHITDKDDFA 600
 
Query: 1959 NNREFITMVVYKTDGKKVYYPADPPPYIDGIRINSPHYLTKIKLTTPGTHTFTLVVSQYE 2138
            NNREFITMVVYKTDGKKVYYPADPPPYIDGIRINSPHYLTKIKLTTPGTHTFTLVVSQYE
Sbjct: 601  NNREFITMVVYKTDGKKVYYPADPPPYIDGIRINSPHYLTKIKLTTPGTHTFTLVVSQYE 660
 
Query: 2139 KQNTIHYTVRVYSACSFTFSKIPSPYTLSKRINGKWSGQSAGGCGNFQETHKNNPIYQFH 2318
            KQNTIHYTVRVYSACSFTFSKIPSPYTLSKRINGKWSGQSAGGCGNFQETHKNNPIYQFH
Sbjct: 661  KQNTIHYTVRVYSACSFTFSKIPSPYTLSKRINGKWSGQSAGGCGNFQETHKNNPIYQFH 720
 
Query: 2319 IDKTGPLLIELRGPRQYSVGFEVVAVSIMGDPGPHGFQRKSSGDYRCGFCYLELENIPAG 2498
            IDKTGPLLIELRGPRQYSVGFEVVAVSIMGDPGPHGFQRKSSGDYRCGFCYLELENIPAG
Sbjct: 721  IDKTGPLLIELRGPRQYSVGFEVVAVSIMGDPGPHGFQRKSSGDYRCGFCYLELENIPAG 780
 
Query: 2499 IFNIIPSTFLPKQEGPFFLDFNSTVPIKTTQLQ 2597
            IFNIIPSTFLPKQEGPFFLDFNSTVPIKTTQLQ
Sbjct: 781  IFNIIPSTFLPKQEGPFFLDFNSTVPIKTTQLQ 813
 
 
 

Construct

Gene:

565

 

GI Number(s):

6753257

ES Cell Line:

4821

 

 

Accession Number:

NM_009796

 

 

Family:

Protease

 

 

Subfamily:

Cysteine

 

 

Species:

Mouse

 

 

 

Sequence Type:

full length cDNA

 

 

 

 

 


Sequence Length:

2848 bp

 

red underlined text = deleted in gene sequence

Sequence Deleted:

from base 1011 to base 1032

 

black text = gene sequence flanking deleted sequence



GTCTGAGGTAGTGGGGCTCCGTCATCTCTTCCGCCGCCCTCGAAAGTAAACTTTGCCACT
CCACCCGCGGGGTCCCCAGTTCTCGCCGCAGTGCCTCCGGGCGTCGGGGTTCCGCGTCAG
GGCCTGTCTGCCTCCCCCGGCGTGGGGCCTTGGGCGCCATGGACGCGTCGGCGCTGGAGC
GGGACGCCGTGCAGTTCGCGCGGCTGGCAGTGCAGCGCGACCACGAAGGCCGCTACTCTG
AGGCGGTGTTTTACTACAAGGAAGCTGCCCAAGCTTTAATTTATGCTGAGATGGCAGGAT
CCAGCCTAGAAAGGATCCAAGAGAAGATCAATGAGTATCTGGAAAGAGTGCAAGCCCTTC
ACTCTGCAGTTCAGTCAAAGAGTACTGATCCTTTGAAATCAAAACACCAGTTGGATCTAG
AACGTGCCCATTTTCTTGTTACACAAGCTTTCGATGAAGATGAAAAGGGGAATGTGGAAG
ATGCCATAGAATTATATACAGAAGCTGTGGAACTCTGTCTAAAAACCTCTTCTGAAACTG
CTGACAAGACTCTGCAAAATAAATTGAAACAGTTGGCTCGACAGGCACTAGATAGAGCAG
AAGCATTGAGTGAACCTTTGACAAAGCCATTTTGCAAACTCAAGTCAGCAAATATGAAAA
CAAAGACTCCTCCAGTAAGAACACACTTTCCTCTAGGACCTAATCCTTTCGTTGAAAAGC
CTCAAGCATTTATAAGTCCACAGTCATGTGATGCGCAAGGACAGAAGTACACAGCAGAAG
AGATAGAAGTGCTCAGGACAACATCAAAAATAAATGGTGTAGAATATGTTCCTTTCATGA
GTGTTGATCTAAGGGAACGCTTTGCTTATCCAATGCCTTTTTGTGATAGGTTGGGCAAGC
TTCCATTATCACCTAAACAAAAAACTACATTTTCTAAATGGGTACGACCGGAAGATCTCA
CCAACAATCCTACAATGATATACACTGTGTCCAGTTTTAGCATAAAGCAG
ACAATAGTAT
CAGATTGTTCCT
TTGTGGCATCGTTGGCTATCAGTGCAGCGTATGAGAGACGATTCAATA
AGAAGTTAATTACCAGCATAATTTACCCCCAGAATAAGGATGGAGAACCAGAGTACAATC
CCTGTGGAAAGTATATGGTAAAGCTTCACCTCAATGGTGTTCCGAGAAAGGTGATAATTG
ATGACCAGTTACCTGTTGATCACAAAGGAGAATTGCTGTGTTCTTATTCCAACAATAAAA
GTGAATTGTGGGTGTCACTTATAGAGAAAGCATACATGAAAGTTATGGGAGGATATGATT
TCCCAGGATCCAACTCAAATATTGATCTGCATGCACTGACTGGGTGGATACCAGAAAGGA
TTGCCATGCATTCAGATAGCCAGACGTTCAGTAAGGATAACTCATTCAGAATGCTTTATC
AAAGATTTCACAAAGGAGATGTCCTCATCACCGCATCGACGGGAGTAATGACTGAAGCTG
AAGGGGAGAAGTGGGGCCTGGTTCCCACTCATGCCTATGCGGTTTTGGATATTAGAGAGT
TCAAGGGGCTGAGATTTATTCAGTTAAAAAACCCTTGGAGCCATTTACGCTGGAAAGGAA
GGTACAGTGAGAATGATGTGAAAAACTGGACCCCAGAATTGCAGAAGTATTTAAACTTTG
ATCCACGAACAGCTCAGAAAATAGATAATGGAATATTTTGGATTTCCTGGGATGATCTTT
GCCAGTATTATGATGTAGTTTATTTGAGTTGGAATCCAGCTCTTTTTAAAGAATCAACTT
GCATTCACAGTACTTGGGATGCTAAACAAGGACCTGTGAAAGATGCTTATAGCCTGGCCA
ACAACCCCCAGTACAAATTGGAGGTGCAGTGTCCACAAGGGGGTGCTGCAGTTTGGGTTT
TGCTTAGTAGACACATAACAGACAAGGATGACTTCGCAAATAACCGTGAGTTTATCACAA
TGGTTGTGTACAAGACTGATGGCAAGAAGGTTTACTACCCAGCTGACCCACCTCCATACA
TTGATGGGATTCGAATTAACAGCCCTCATTACTTGACTAAGATAAAGCTGACCACGCCCG
GGACTCACACCTTCACCCTAGTGGTTTCTCAGTATGAGAAACAGAATACGATTCATTACA
CAGTCCGGGTGTATTCAGCCTGCAGCTTTACTTTTTCGAAGATCCCTTCACCATACACCT
TATCCAAACGGATAAATGGAAAGTGGAGTGGTCAAAGTGCTGGAGGATGTGGAAATTTTC
AGGAGACTCACAAAAATAACCCCATCTACCAATTCCATATAGATAAGACTGGACCGTTAT
TGATTGAGCTGAGAGGACCAAGGCAATACAGTGTTGGATTTGAGGTTGTAGCAGTTTCCA
TAATGGGAGATCCTGGTCCTCATGGCTTTCAGAGGAAATCTAGTGGTGACTATAGGTGTG
GATTTTGCTACTTAGAACTGGAAAACATACCTGCTGGGATCTTTAATATCATCCCTAGTA
CTTTTTTGCCTAAACAAGAAGGACCTTTTTTCTTGGACTTTAATAGTACTGTCCCCATCA
AGACAACACAACTCCAATGATTGAGAACTCTCAGCTATTACTGGATTCCATATTTCCCAG
TCTCAGCTCTTCAGACAAGGAGACAAATCACGATGGTGAACCATCAGAATGGAATTAAAT
CAAAAGCATGTTACGGAGTCTGGTGCTCGTTGTGGTGTTTGGTAAGGATTTTAGATAGTG
GAATTACCTTAATCACCTGAAATACTGTTTACATGGTTTGTGCACACAGAGCTGGCGTGT
ACTCGGTGGCGGCTCTGTGTAAATCACA




Targeting Vector* (genomic sequence)

    LacZ-Neo

Construct Number:  9677

Arm Length:
5':  5  kb
3':  1.6  kb


  * Not drawn to scale



 

5 ' PCR Reconfirmation

Gene:

565

ES Line:

4821

Gene Specific Oligo (GS):
AGTAAGGAGGCGCTGGTTGATGATTG

34285



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

34285 + NEO1 (11193)

5125

3

34285 + NEO2 (11192)

5135

4

34285 + NEO3 (2416)

5700

5

DNA Sample Control

5167

6

Positive Control

5000

 


 

3 ' PCR Reconfirmation

Gene:

565

ES Line:

4821

Gene Specific Oligo (GS):
TTGAGACAGGGTTTCTCTGCGTAGTC

34284



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

34284 + NEO1 (1431)

2900

3

34284 + NEO2 (1432)

2850

4

34284 + NEO3 (1170)

2150

5

DNA Sample Control

5167

6

Positive Control

1900

 


 

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:

565

ES Line:

4821

Construct:

9677

Blot Number:

4424

Gene Side:

5'

 

 

MW Marker

Lane

ES Line

Enzyme

Yes

1

 

 

 

2

Parent 1

Kpn I

 

3

Parent 2

Kpn I

 

4

4821

Kpn I

 

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 #565 allele.  In contrast, the ES#4821  line showed an additional band representing the targeted allele from the expected homologous recombination event.  
Note: All lanes represent data from the same blot.

 

Genotyping: Gene 565 (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)

56839

TCCTCTCTACTCTCACATGCATAAG

400

NEO(T)

306

TCAGCAGCCTCTGTTCCACATACACTTC

400

GS(E1)

56838

TCTGTGTATCCATACTCATGTGTAC

400

 

D) Reaction Conditions:

Allele

Expected Size

Oligos to use

Targeted

299

oligo 306 + oligo 56839

Endogenous

630

oligo 56838 + oligo 56839