Abstract:
This invention relates to identification of cancer genes using a high-throughput forward genetic screening method called insertion mutation scanning (IMS).

Description:
FIELD OF THE INVENTION  
       [0001]     This invention relates to identification of cancer genes using a high-throughput forward genetic screening method called insertion mutation scanning (IMS).  
       BRIEF SUMMARY OF THE INVENTION  
       [0002]     The invention includes a highly efficient and scalable method for large scale isolation of host/provirus fragments. The invention is an anchored-PCR method designed to amplify host/virus junction fragments from new clonally integrated proviruses present in tumors induced by certain retroviruses or pathogenic endogenous proviruses. These proviruses represent the somatic mutations which caused the tumors from which the integrants were isolated. The IMS technique has utility with respect to a cancer gene discovery strategy called provirus tagging (see, e.g., US 2004/0142336A1). Because of the efficiency and scalability of the IMS procedure, it is useful technology that makes a saturation screen of the cancer genome feasible. The IMS procedure implements an anchored-PCR approach to provirus tagging. Advantages over previous methods include improvements in (1) sensitivity, (2) signal-to-noise ratio, (3) efficiency of mutation recovery, and (4) scalability. The invention includes nucleotide sequences (both complete sequences and host sequences) shown in Table 1. 
     
    
     BRIEF DESCRIPTION OF THE DRAWINGS  
       [0003]      FIG. 1  is a schematic of bioinformatics analysis.  
         [0004]      FIG. 2  shows the amplification results from 24 independent WXG/WLC tumors induced by MMTV. The upper common band contains the env region flanking the 3′ LTR and the lower common band contains the host sequence flanking the endogenous Mtv2 provirus which is common to all the tumors in the panel. Each unique band contains the 5′ host flanking sequence from a unique provirus insertion mutation that tags a cancer gene. 
     
    
     DETAILED DESCRIPTION  
       [0005]     The methods of the invention can be used to prepare nucleotide sequences from cancer genes. A sample protocol illustrating the methods of the invention is as follows.  
         [0006]     1. Extract genomic DNA from frozen tumor tissues using a SNAP-O-SOL kit from Biotecx (Houston, Tex.), according to the manufacturer&#39;s instructions.  
         [0007]     2. Digest 750 ng of genomic DNA with 10 units of NlaIII from New England BioLabs (Beverly, Mass.) in a total of 20 μl for one hour at 37° C. in the manufacturer&#39;s buffer (NEBuffer 4 with supplemental BSA).  
         [0008]     3. Heat inactivate the enzyme at 65° C. for 25 min and place onto ice.  
         [0009]     4. Ligate anchors onto 150 ng of NlaIII digested DNA (4 μl). Anchors consist of two annealed oligos with a 3′ CATG overhang. The oligos, synthesized by Integrated DNA Technologies (Coralville, Iowa), have the following sequence: 
    n0131 5′-CCTGA CTATG GAGTT CACTG CGTGC TACAC TATCA GTAGG CATG -3′    n0157 5′-CCTAC TGATA GTGTA GCACG-N-3′    note: n0157 contains a 3′ amino modifier (with a C7 spacer).     Prior to use, dilute oligos from a concentrated stock solution to 4 μM each in TE.     Mix equal parts to make a 2 μM solution.     Heat to 95° C. for 2 min and lower temperature to 45° C. for 10 min.    
 
         [0016]     5. Ligation:  
         [0017]     DNA Ligation Kit Version 1 from Takara Shuzo Co., Ltd. (Otsu; Shiga, Japan) 
    4 μl NlaIII digested DNA     1 μl 2 μM n0131/n0157     25 μl Solution A     05 μl Solution B     Mix all components on ice. Incubate at 16° C. for 1 hour.     Purify using Qiaquick PCR Purification Kit from Qiagen, Inc. (Santa Clara, Calif.) according to the manufacturer&#39;s instructions. Elute into 50 μl 10 mM Tris-HCl, pH 8.5 and place on ice.    
 
         [0024]     6. Amplify the ligated material through two successive rounds of PCR using nested primers as follows.  
         [0025]     7. PCR reaction 1: 
    5 μl anchor ligated DNA     1X Qiagen PCR Buffer with 1.5 mM MgCl2 (Qiagen; Santa Clara, Calif.)     200 μM dNTPs     50 U/ml HotStarTaq DNA Polymerase (Qiagen; Santa Clara, Calif.)     0.2 U Perfect Match PCR Enhancer (Stratagene; LaJolla, Calif.)     0.2 μM of each of the amplification primers     n0121 5′-CCTGA CTATG GAGTT CACTG-3′    n0138 5′-GATGG GCTCA TCCGT TTGTG-3′    H20 to a 50 μl reaction volume.     Perform the reactions in an Amplitron II thermal cycler from Thermolyne (Dubuque, Iowa) using a step down PCR reaction with denaturation, annealing, and extension temperatures of 94° C., 60-56° C., and 72° C., respectively. During the first 5 cycles, drop the annealing temperature in one degree increments from 60° C. to 56° C. Perform the last 60 cycles at a constant annealing temperature of 56° C., for a total of 65 cycles during the first round.     Cycling conditions:     95° C., 15 min dwell     2 cycles (94° C., 30 sec; 60° C., 30 sec; 72° C., 30 sec)     1 cycle (94° C., 30 sec; 59° C., 30 sec; 72° C., 30 sec)     1 cycle (94° C., 30 sec; 58° C., 30 sec; 72° C., 30 sec)     1 cycle (94° C., 30 sec; 57° C., 30 sec; 72° C., 30 sec)     60 cycles (94° C., 30 sec; 56° C., 30 sec; 72° C., 30 sec)     72° C., 10 min dwell    
 
         [0044]     8. PCR reaction 2:  
         [0045]     9. Shotgun clone 1 μl of this PCR product using a TA TOPO Cloning Kit from Invitrogen (Carlsbad, Calif.), according to the manufacturer&#39;s instructions with the following exception. Do not incubated transformed cells in a shaker after heat shock and subsequent ice step. Plate directly onto prewarmed LB Amp100/Kan 5 plates to ensure more accurate cloning representation.  
         [0046]     10. PCR screen white colonies for insert size as described below.  
         [0047]     11. Grown all screened colonies overnight in 2 ml LB Amp100/Kan50.  
         [0048]     12. Miniprep cultures with correct insert sized using a QIAprep Miniprep Kit from Qiagen (Santa Clara, Calif.).  
         [0049]     13. Send plasmids to University of California, Davis DBS Automated Sequencing Facility for insert sequence determination using PRISM sequencers, reagents, and software from ABI (Foster City, Calif.).  
         [0050]     14. PCR screening for insert size: 
    Pick 24 white colonies/tumor into 20 μl LB Amp100/Kan50.     PCR reaction:     1 μl culture     1X Qiagen PCR Buffer with 1.5 mM MgCl2 (Santa Clara, Calif.)     200 μM dNTPs     20 U/ml Qiagen Taq Polymerase     0.2 μM each of the amplification primers     n0122 5′-CGTGC TACAC TATCA GTAGG-3′    n0036 5′-CAACC CCTTG GCTGC TTCTC-3′    50 μl reaction volume.    
 
         [0061]     Perform the reactions in an Amplitron II thermal cycler from Thermolyne (Dubuque, Iowa) for 30 cycles with denaturation, annealing, and extension temperatures of 94° C., 56° C., and 72° C., respectively.  
                                                                           TABLE 1                           IST Sequences                            INSERT   CANCER           TAG   BAND   COMPLETE SEQUENCE   HOST SEQUENCE   LENGTH   GENE                    IST000001   MM0001B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCAT   CATGAAGACCCCAGGCTCAGTT   264   unknown                   GAAGACCCCAGGCTCAGTTTCTGCTCCCAGGAGAA   TCTGCTCCCAGGAGAAAGTCCT               AGTCCTGCTCTCTAGTCTGCCCTGAGGGCCAAACA   GCTCTCTAGTCTGCCCTGAGGG               CGGGACTCTTGGTGCACCTCGTCAGTTTTGTGCAG   CCAAACACGGGACTCTTGGTGC               TGCTTTAGCAGGGCCAGAGAGCCCATCTAGTGGCC   ACCTCGTCCAGTTTTGTGCAGT               AGGGCAGAACCCTGTCCCTGGCTGCATCTGGTCTT   GCTTTAGCAGGGCCAGAAGCCC               CAGCTCAGGAATGTGAGATTGTCTTGTTAGTTTTT   ATCTAGTGGCCAGGGCAGAACC               GAAGTAACACAGTTCCTGCCTAGCTCCAAGAGTGG   CTGTCCCTGGCTGCATCTGGTC               CGCTCCCTCCCACTTGCCGCGCCTGCAGCAGAAAT   TTCAGCTCAGGAATGTGAGATT               GGTTGAACTCCCGAGAGTGTCCTACACTTAGGGGA   GTCTTGTTAGTTTTTGAAGTAA               GAAGCAGCCAAGGGGTTGAAGGGCGAATTC   CAACAGTTCCTGCCTAGCTCCA                   AGAGTGGCGCTCCTCCCACT{circumflex over ( )}               IST000002   MM0001C   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCAT   CATGCTCATAATTCTGCAGTGC   156   unknown               GCTCATAATTCTGCAGTGCCTTCTCATAACACAGG   CTTCTCATAACACAGGATAAAA               ATAAAACACTCTAACCTTTAACATTATACTTGAAA   CACTCTAACCTTTAACATTATA               ACTTATGTGGTTTTTTCCTACCGAGTCATATCAAA   CTTGAAAACTTATGTGGTTTTT               CCAGTCTCCCTCTCCACTCACAAGGATCCAGTCAC   TCCTACCAGAGTCATATCAAAC               AATGGCCTTTTATGCCGCGCCTGCAGCAGAAATGG   CAGTCTCCCTCTCCACTACAAG               TTGAACTCCCGAGAGTGTCCTACACTTAGGGGAGA   GATCCAGTCACAATGGCCTTTT               AGCAGCCAAGGGTTGAAGGGCGAATTC   A{circumflex over ( )}               IST000003   MM0001D   GAATTCGCCCTCGTGCTACACTATCAGTAGGCAT   CATGTCCTACAGTGAACATTTC   130   unknown               GTCCTACAGTGAACATTTCTATATTTTCCACTTTT   TATATTTTCCACTTTTTTCAGT               TTCAGTTTTCCTCGCCATATTTCACGTCCTAAAGT   TTTCCTCGCCATATTTCACGTC               GTGTATTTCTCATTTTCCGTGATTTCAGTTTTCTC   CTAAAGTGTGTATTTCTCATTT               GCCATATTCCAGGTCCTACAGTGCCGCGCCTGCAG   TCCGTGATTTTCAGTTTTCTCG               CAGAAATGGTTGAACTCCCGAGAGTGTCCTACACT   CCATATTCCAGGTCCTAAG{circumflex over ( )}               TAGGGGAGAAGCAGCCAAGGGGTTGAAGGGCGAAT               TC               IST000004   MM0001E   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCAT   CATGGCGAGATTCTGTGTCCAA   101   Wnt3               GGCGAGATTCTGTGTCCAAGCTGCCTCTACTCGTG   GCTGCCTCTACTCGTGACATTC               ACATTCCAAGATGCCTCTGAGGTGGGAACTGTGAA   CAAGATGCCTCTGAGGTGGGAA               ATAGGAGAGAGCCCCACAGTCCCTCTTTGCCGCGC   CTGTGAAATAGGACAGAGCCCC               CTACAACAAAAATGGTTGAACTCCCGAGAGTGTCC   ACAGTCCCCTCTT{circumflex over ( )}               TACACTTAGGGGAGAAGCAGCCAAGGGGTTGAAGG               GCGAATTC               IST000005   MM0007C   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCAT   CATGTGTTTCCCGATTTTAGTT   89   unknown               GTGTTTCCCGATTTTAGTTGGATTTTTTTTCTCCC   GGATTTTTTTTCTCCCAGGTTT               AGGTTTCTGCAGTGTCCCCACCCCCCACCCCCATT   CTGCAGTGTCCCCACCCCCCAC               TTCTCCCACTCTTACCTGCCGCCCTGCAGCAGAAA   CCCCATTTTCTCCCACTCTTAC               TGGTTGAACTCCCGAGAGTGTCCTACACTTAGGGG   C{circumflex over ( )}               AGAAGCAGCCAAGGGGTTGAAGGGCGAATTC               IST000006   MM0014B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCAT   CATGGTGACAGTCCCTGAGTAA   106   unknown               GGTGACAGTCCCTGAGTAAAGGCTGGCTACTCCAT   AGGCTGGCTACTCCATCTTAGT               CTTAGTGGATTTGTTTTTATAAAGAAGGCAGACGA   GGATTTGTTTTTATAAAGAAGG               GAACCAGAAATAACAACACATTTAAGCGTATGTGC   CAGACGAGAACCAGAAATAACA               CGCGCCTGCAGCAGAAATGGTTGAACTCCCGAGAG   ACACATTGTAAGCGTATG{circumflex over ( )}               TGTCCTACACTTAGGGGAGAAGCAGCCAAGGGGTT               GAAGGGCGAATTC               IST000007   MM0008B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCAT   CATGTATTTTTGCTGGGACTGT   50   unknown               GTATTTTTGCTGGGACTGTTCCCTCTGGGAACTGC   TCCCTCTGGGAACTGCGGGATG               GGGATGGGGGTGTGCCGCGCCTGCAGCAGAAATGG   GGGGTG{circumflex over ( )}               TTGAACTCCCGAGAGTGTCCTAACTTAGGGGAGAA               GCAGCCAAGGGGTTGAAGGGCGAATTC               IST000008   MM0003B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATG   CATGGCAAGATGGAGACTTTG   75   Fgf3               GCAAGATGGAGACTTTGTCTACCAGGGCCACTCCAA   TCTACCAGGGCCACTCCAAGC               GCACCCAGCTGCATACAGGTGGACTGGCTGTGGCCT   ACCCAGCTGCATACAGGTGGA               GCCGCGCCTGCAGCAGAAAGGTTGAACTCCCGAGAG   CTGGCTGTGGCC{circumflex over ( )}               TGTCCTACACTTAGGGGAGAAGCAGCCAAGGGGTTG               AAGGGCGAATTC               IST000009   MM0010B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATG   CATGTAAGAAAAACAAGATCA   170   unknown               TAAGAAAAACAAGATCAAAGAAATGAGTCAAAAAGG   AAGAAATGAGTCAAAAAGGCT               CTCTAACAGGGCTTGAAGGTGAGGTAGAAATGACAG   CTAACAGGGCTTGAAGGTGAG               AAGACATAGAAATCCAATATCAGAAGGCAGAGTTTG   GTAGAAATGACAGAAGACATA               TATGGGACAGATACTCTCTATATATATGCTAGCACA   GAAATCCAATATTCAGAAGGC               CCTACTACTCAGCTGTTTCTCTGCCGCGCCTGCAGC   AGAGTTTGTATGGGACAGATA               AGAAATGGTTGAACTCCCAAGAGTGTTCTACACTTA   CCTCTATATATATGCTAGCAC               GGGAGAAGCAGCCAAGGGGTTGAAGGGCGAATTC   ACCTACTACTCAGCTGTTTCT                   C{circumflex over ( )}               IST000010   MM0015B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATG   CATGTCCTACAGTGGATATTT   135   unknown               TCCTACAGTGGATATTTCTAAATTTTCCTCCTTTTT   CTAAATTTTCCTCCTTTTTCA               CAGTTTTCCTCGCCATATTTGAAGTCCTAAAGTGTG   GTTTTCCTCGCCATATTTGAA               TATTTCTCATATCCTGTGATTTCAGTTTTCTCSCCA   GTCCTAAAGTGTGTATTTCTC               TATTCCAGGTCCTACAGTGTGCTGCCGC0CCTGCAG   ATATCCTGTGATTTTCAGTTT               CAGAAATGGTTGAACTCCCGAGAGTGTCCTACACTT   TCTCGCCATATTCCAGGTCCT               AGGGGAGAAGCAGCCAAGGGGTCGAAGGGCGAATTC   AAGTGTGC{circumflex over ( )}               IST000011   MM0015C   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATG   CATGTGGAGGCCAGAAGTCAA   59   unknown               TGGAGGCCAGAAGTCAACATATAGTCTCCTTCCCAA   CATATAGTCTCCTTCCCAATT               TTACTTGTCACTGGAGAGCTGCCGCGCCTGCAGCAG   ACTTGTCACTGGAGAGC{circumflex over ( )}               AAATGGTTGAACTCCCGAGGTGTCCTACACTTAGGG               GAGAAGCAGCCAAGGGGTTGAAGGGCGAATTC               IST000012   MM0013B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATG   CATGCTACATTTTCATATTGT   56   unknown               CTACATTTTCATATTGTGTGCATACAAGGTTTCCCT   GTGCATACAAGGTTTCCCTTT               TTATTATGTGGGGTAATGCCGCGCCTGCAGCAGAAA   ATTATGTGGGGTAA{circumflex over ( )}               TGGTTGAACTCCCGAGAGTTCCTACACTTAGGGGAG               AAGCAGCCAAGGGGTTGAAGGGCGAATTC               IST000013   MM0013B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGAATG   AATGGGAGGAAGGACTGTGGA   43   unknown               GGAGGAAGGACTGTGGAAGGCAGAGGTGGAAATATC   AGGCAGAGGTGGAAATATCTG               TGCTGCCGCGCCTGCAGCAGAAATGGTTGAACTCCC   C{circumflex over ( )}               GAGAGTGTCCTACACTTAGGGAGAAGCAGCCAAGGG               GTTGAAGGGCGAATTC               IST000014   MM0006B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATG   CATGCTGGCTGTTCCTGCAGC   46   Fgf8               CTGGCTGTTCCTGCAGCCCAGCTACTAGGACAATCT   CCAGCTACTAGGACAATCTGG               GGAAACTGCCGCGCCTGCAGCAGAAATGGTTGAACT   AAAC{circumflex over ( )}               CCCGAGAGTGTCCTACACTAGGGGAGAAGCAGCCAA               GGGGTTGAAGGGCGAATTC               IST000015   MM0007B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATG   CATGGAAAGATAATGTGTAAA   244   unknown               GAAAGATAATGTGTAAATTTGGGTTTGCCGTGGAAA   TTTGGGTTTGCCGTGGAAAAC               ACTTTGGTTTCTCCATCAATGGTAATTGAGAGTTTG   TTTGGTTTCTCCATCAATGGT               GCTGGGTATAGTAGCCTGGCTGGCATTTTTGTTCTC   AATTGAGAGTTTGGCTGGGTA               TTAAGGTCTGTATAAAGTCTGTCCAGGATCTTCTGA   TAGTAGCCTGGGCTGGCATTT               CTTTCATAATGTCTGGTGTAAAGTCTGGTGTAATTC   TTGTTCTCTTAAGGTCTGTAT               TGACAGGCCTGCCTTTATATGTTACTTGACCTTTTT   AAGTCTGTCCAGGATCTTCTG               CCTTACTGCTTTTAATATTCTATGCCGCGCCTGCAG   ACTTTCATAATGTCTGGTGTA               CAGAAATGGTTGAACTCCCGAGAGTGTCCTACACTT   AAGTCTGGTGTAATTCTGACA               AGGGGAGAAGCAGCCAAGGGGTTGAAGGGCGAATTC   GGCCTGCCTTTATATGTTACT                   TGACCTTTTTCCCTTACTGCT                   TTTAATATTCTA{circumflex over ( )}               IST000016   MM0010C   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATG   CATGGGTCCCTGAAGGGTCTC   79   unknown               GGTCCCTGAAGGGTCTCTCCTTTAGCAAACCCCTGT   TCCTTTAGCAAACCCCTGTAC               ACAGTTGAAGTGATTTTTCAGGTACCCATTGGTCTT   AGTTGAAGTGATTTTTCAGGT               AGCTGCCGCGCCTGCAGCAAAATGGTTGAACTCCCG   ACCCATTGGTCTTAGC{circumflex over ( )}               AGAGTGTCCTACACTTAGGGGAGAAGCAGCCAAGGG               GTTGAAGGGCGAATTC               IST000017   MM0004B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCA   CATGTTGTCCTTTGAGCATCTC   127   unknown               TGTTGTCCTTTGAGCATCTCCACCTAGCAGCTGA   CACCTAGCAGCTGACTTAGACA               CTTAGACAGATACACACACCCACAGGTGAACAGT   GATACACACACCCACAGGTGAA               GGATGGAGATTGGGGACTCTTATGGAGAATGGGA   CAGTGGATGGAGATTGGGGACT               GGAAGGATTTTGGGCCCTAAGGTGCCGCGCCTGC   CTTATGGAAGAATGGGAGGAAG               AGCAGAAATGGTTGAACTCCCGAGAGTGTCCTAC   GATTTTGGGCCCTAAGG{circumflex over ( )}               ACTTANGGGAGAAGCAACCAAGGGGTGAAGGGCG               AATTC               IST000018   MM0004C   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCA   CATGTGACACTGGCTTTGGAGT   81   unknown               TGTGACACTGGCTTTGGAGTCATACATAGAAGGG   CATACATAGAAGGGGTTGGGAC               GTTGGGACAATTGATGCTAGTTAGCTAGAGCTAA   AATTGATGCTAGTTAGCTAGAG               GAATATAGCAGTGCCGCGCCTGCAGAGAAATGGT   CTAAGAATATAGCAG{circumflex over ( )}               TGAACTCCCGAGAGTGTCCTACACTTAGGGGAGA               AGCAGCCAAGGGGTTTAAGGGCGAATTC               IST000019   MM0005B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCA   CATGGTCTCTAGAGCTAAGAGA   173   unknown               TGGTCTCTAGAGCTAAGAGATACCAATGCTGCGG   TACCAATGCTGCGGCAGGCAGT               CAGGCAGTTTTTATTACAATCATTACAGTTTTGA   TTTTATTACAATCATTACAGTT               CAGCGTCTGGCCGTGTGCCAAGGCTGCCTTCATC   TTGACAGCGTCTGGCCGTGTGC               CCTGAGCTCGGTGATGCTTCTGTCCTGGTCTTCT   CAAGGCTGGCCTTCATCCCTGA               GGCTCATCACAGCTTAAGAAAGTAGCTGCTTCTC   GCTCGGTGATGCTTCTGCCTGG               TGCCGCGCCTGCAGCAGAAATGGTTGAACTCCCG   TCTTCTGGCTCATCACAGCTTA               AGAGTGTCCTACACTTGGGGAGAAGCAGCCAAGG   AGAAAGTAGCTGCTTCTC{circumflex over ( )}               GGTTGAAGGGCGAATTC               IST000020   MM0002C   GAATTCGCCCTTCGTGCTACACTATCAGTAGGGA   GATGGAACACTCAAAGCTGGCC   115   unknown               TGGAACACTCAAAGCTGGCCAGGGCCCATTTACC   AGGGCCCATTTACCAGGTATCC               AGGTATCCTTTGCCTTCTCAGCTGATGGGCATCA   TTTGCCTTCTCAGCTGATGGGC               ACACATTAATTCACATATGACTCGTTGTGTCATA   ATCAACACATTAATTCACATAT               TCAATAGTATTGCCGCGCCTGCAGCAGAAATGGT   GACTCGTTTGTGTCATATCAAT               TGAACTCCCGAGAGTGTCCTACACTTAGGGGAGA   AGTAT{circumflex over ( )}               AGCAGCCAAGGGGTTGTTTCCCACCAAGGACGAC               CCGTCTGCGCACAAACGATGAGCCCATCAAGGGC               GAATTC               IST000021   MM0002B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCA   CATGGGGGGGGGTACCAAGAAG   271   unknown               TGGGGGGGGGTACCCAAGAAGGGACTGCTGCGAT   GGACTGCTGCGATTGGGATGTA               TGGGATGTAAATAAATAAATAAATAGAATAAACA   AATAAATAAATAAATAGAATAA               AAACCCAAAAACAAACAGAAACCTAACTCAATAA   ACAAAACCCAAAAACAAACAGA               CTGCAGAAATGACTCTTGCTCTTTTCTGGTAAGG   AACCTAAACTCAATAACTGCAG               TTAGAAGCAGGTTACAAATCTATATTAGAGATGG   AAATGACTCTTGCTCTTTCTGG               AGGCATTTCACACCAGCATAGGTATAGGAAGTAG   TAAGGTTAGAAGCAGGTTACAA               ATGAAATGAGGACTACCTAGAGTCTGTTTGTCAC   ATCTATATTAGAGATGGAGGCA               AACCAATTCTGAGTGATTTCACTGAGATATTGCC   TTTCACACCAGCATAGGTATAG               GCGCCTGCAGCAGAAATGGTTGAACTCCCGAGAG   GAAGTAGATGAAATGAGGACTA               TGTCCTACACTTAGGGGAGAAGCAGCCAAGGGGT   CACTAGAGTCTGTTTGTCACAA               TGAAGGGGAATTC   CCAATTCTGAGTATTTCACTGA                   GATAT{circumflex over ( )}               IST000022   MM0016D   GAATTCGCCCTTCGTGCTACACTATCAGTAGGGA   GATGTACTGAATCCCTGAAGTT   66   unknown               TGTACTGAATCCCTGAAGTTGATGCTGAGCACCA   GATGCTGAGCACCATCTTGTGC               TCTTGTGCTGTTCTACCGCATTTACTGGGGTGCC   TGTTCTACCGCATTTACTGGGG{circumflex over ( )}               GCGCCTGCAGCAGAAATGGTTGAACCCCGAGAGT               GTCCTACACTTAGGGGAGAAGCAGCCAAGGGGTT               GAAGGGCGAATTC               IST000023   MM0016B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCA   CATGAGTACATTTCATTTTTTC   212   unknown               TGAGTACATTTCATTTTTTCCCACAAACTGCACA   CCACAAACTGCACAGACCAACT               GACCAACTTTGAGAAATGCCTGTTTATGAATGAC   TTGAGAAATGCCTGTTTATGAA               CAGAGATCACGTCTTCCATTTTTAACACTTAGAG   TGACCAGAGATCACGTCTTCCA               ATGTGGAAGACTTCTGGTTAGCCCACTTATCCCT   TTTTTAATCACTTAGAGATGTG               TCCCAGACTAGCAGTCAAGGCCAAGCAAAGGGAG   GAAGACTTCTGGTTAGCCACTT               AATTTGGGCTGGTCCTATCCCCCCTGCTGGCTAC   ATCCCTTCCCAGACTAGCAGTC               TGAACTGCCGCGCCTGAGCAGAAATGGTTGAACT   AAGGCCAAGCAAAGGGAGAATT               CCCGAGGGTGTCCTACACTTAGGGGAGAAGCAGC   TGGGCTGGTCCTATCCCCCCTG               CAAGGGTTGTTTCCCACCAAGGACGACCCGTCTG   CTGGCTACTGAAC{circumflex over ( )}               CGCACAAACGGATGAGCCCATCAAGGGCGAATTC               IST000024   MM0016C   GAATTCGCCCTTCGTGCTACACTATCAGTAGG   CATGGATTATCTCTCTTCTATCCTT   125   unknown               CATGGATTATCTCTCTTCTATCCTTCCATTTT   CCATTTTTGTTTTTGTTTGTTGGGT               TGTTTTTGTTTGTTGGGTTGGTTTTGATTTTT   TGGTTTTGATTTTTGAGGCAGGGTT               GAGGCAGGGTTTCCCTGTGCATCCCTGCCTGC   TCCCTGTGCATCCCTGCCTGGCTAT               TATCCTCGAACTTGACCTCACTCTGTAGCGCC   CCTCGAACTTGACCTCACTCTGTAG{circumflex over ( )}               GCGCCTGCAGCAGAAATGGTTGAACTCCCGAG               AGTGTCCTACACTTAGGGGAGAAGCAGCCAAG               GGGTTGAAGGGCGAATTC               IST000025   MM0009C   GAATTCGCCCTTCGTGCTACACTATCAGTAGG   CATGTAATGCACAATATATGAGTCC   96   unknown               CATGTAATGCACAATATATGAGTCCTGATATT   TGATATTTATCCCTTACAGTCAATA               TATCCCTTACAGTCAATATAGCTTTGTCTACA   TAGCTTTGTCTACAAATTTTTGACA               AATTTTTGACATAAAGTAGGGCTATTTTTCAT   TAAAGTAGGGCTATTTTTCAA{circumflex over ( )}               GCCGCGCCTGCAGCAGAAATGGTTGAACTCCC               GAGAGTGTCCTACACTTAGGGGAGAAGCAGCC               AAGGGGTTGAAGGGCGAATTC               IST000026   MM0009B   GAATTCGCCCTTCGTGCTACACTATCAGTAGG   CATGGAAATGCAATGAAAGCACACA   301   Unknown               CATGGAAATGCAATGAAAGCACACAATATTGT   ATATTGTTGAACCAAACAGAAAGCT               TGAACCAAACAGAAAGCTCAAAATTAGGCACA   CAAAACTAGGCACAGAAAAGAGATA               GAAAAGAGATACAAACACAAATCTGAACAAAT   CAAACACAAATCTGAACAAATTGAC               GACCTTCTCCCTATAGCATAACTAATATCTCA   CTTCTCCCTATAGCATAACTAATAT               GAGATAAAAGTGGTCTTTATATACCAGGGCGA   CTAGAGATAAAAGTGGTCTTTATAT               AAGAGGTCTAAAAAGAGAGAAATAAAAAATAT   ACCAGGGCGAAAGAGGTCTAAAAAG               GGCATATTTCCTGTCATATGCAGAACCTATTG   AGAGAAATAAAAAATATGGCATATT               AGTCTTTTTGTTTGTTTCTTTCAATACAGCCT   TCCTGTCATATGCAGAACCTATATG               ATGTAGCTCTAGCTGTCCTAGAACTTACTTTG   AGTCTTTTTGTTTGTTTCTTTCAAT               TAGACCAGGCTTGCCGCGCCTGCAGCAGAAAT   ACAGCTATGTAGCTCTAGCTGTCCT               GGTTGAACTCCCGAGAGTGTCCTACACTTGGG   AGAACTTACTTTGTAGACCAGGCT{circumflex over ( )}               GAGAAGCAGCCAAGGGGTTGAAGGGCGAATTC               IST000027   MM0011B   GAATTCGCCCTTCGTGCTACACTATCAGTAGG   CATGTTGACAGTCCTAGAGCAAGAC   163   unknown               CATGTTGACAGTCCTAGAGCAAGACCTGAGGT   CTGAGGTAGAAGAATGGCCTTTAGA               AGAAGAATGGCCTTTAGAAAGTAGGGACTTCA   AAGTAGGGACTTCAGGTGCCTGAAA               GGTGCCTGAAAAATGGAGGGGTGCCAGGGCAA   AATGGAGGGGTGCCAGGGCAGATGT               TGTGGGGAGCCCTGTGAGGAGTGGGGTGTCCC   GGGGAGCCCTGTGAGGAGTGGGGTG               AGGGCAGATGTGGGGAGCCCTGTGAGGAGTGG   TCCAGGGCAGATGTGGGGAGCCCTG               GGTGCCGCGCCTGCAGCAGAAATGGTTGAACT   TGAGGAGTGGGG{circumflex over ( )}               CCCGAGAGTGTCCTACACTTAGGGGAGAAGAG               CCAAGGGGTTGAAGGGCGAATTC               IST000028   MM0011C   GAATTCGCCCTTCGTGCTACACTATCAGTAGG   GATGTCAGAGCCGCAGGAATGCATA   90   unknown               GATGTCAGAGCCGCAGGAATGCATAATGGTCT   ATGGTCTTCTTTTTGTTTGTTTGTT               TCTTTTTGTTTGTTTGTTTTTTGAGACAGGGT   TTTTGAGACAGGGTTTCTCTGTGTA               TTCTCTGTGTAGCCCTGGCTGTCCTGTGCCGG   GCCCTGGCTGTCCTG{circumflex over ( )}               CCTGCAGCAGAAATGGTTGAACTCCCGAGAGT               GTCCTACACTTAGGGGAGAAGCAGCCAAGGGG               TTGAAGGGCGAATTC               IST000029   MM0011D   GAATTCGCCCTTGGTGCTACACTATCAGTAGG   CATGCAGGAGGCCCTCTGTCCTTAA   40   unknown               CATGCAGGAGGCCCTCTGTCCTTAACACAGAA   CACAGAATGTACAGG{circumflex over ( )}               TGTACAGGTGCCGCGCCTGCAGCAGAAATGGT               TGAACTCCCGAGGGTGTCCTACACTTAGGGGG               AAGCAGCCAAGGGGTTGTTTCCCACCAAGGAC               GACCCGTCTGCGCACAAACGGATGAGCCCATC               AAGGGCGAATTC               IST000030   MM0010D   GAATTCGCCCTTCGTGCTACACTATCAGTAGG   GATGCCCGATCAGTTGCTTCTAGAC   48   unknown               GATGCCCGATCAGTTGCTTCTAGACATAAAGA   ATAAAGAAAAATTCGTTGTTTTC{circumflex over ( )}               AAAATTCGTTGTTTTCTGCCGCGCCTGCAGCA               GAAATGGTTGAACTCCCGAGAGTGTCCTACAT               TAGGGGAGAAGCAGCCAAGGGGTTGAAGGGCG               AATTC               IST000031   MM0012B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATGTG   CATGTGAGTTCTTTTGTG   187   unknown               AGTTCTTTTGTGATTGGGTTACCTCACTCAGGATGATA   ATTGGGTTACCTCACTCA               CCCTCCAGATCCATCCATTTGCCTAGGAATGTCATAAA   GGATGATACCCTCCAGAT               TTCATTCTTTTTATAGCTGAGTAGTACTCCATTGTGTA   CCATCCATTTGCCTAGGA               AATGTACCACACTTTCTGTATCCATTCCTCTGTTGAGG   ATGTCATAAATTCATTCT               GGCATCTGGGTTCTTTCCAGCTTCTGGCTGCCGCGCCT   TTTTAATAGCTGAGTAGT               GCAGCAGAAATGGTTGAACTCCCGAGGTGTCCTACACT   ACTCCATTGTGTAAATGT               TAGGGGAGAAGCAGCCAAGGGGTAGAAGGGCGAATTC   ACACACTTTCTGTATCCA                   TTCCTCTGTTGAGGGGCA                   TCTGGGTTCTTTCCAGCT                   TCTGGC{circumflex over ( )}               IST000032   MM0014D   GAATTCGCCCTTCGTGCTACACTATCAGTAGGGATGAT   GATGATG{circumflex over ( )}   7   too short               GTGCCGCGCCTGCAGCAGAAATGGTTGAACTCCCGAGA               GTGTCCTACACTTAGGGGAGAAGCAGCCAAGGGGTTGA               AGGGCGAATTC               IST000033   MM0018B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATGGA   CATGGATCCTGACAACAG   223   unknown               TCCTGACAACAGGCTGAGTAAATGAAGCTGTTTATTCT   GCTGAGTAAATGAAGCTG               CAAGTGCTGAGGGCTGACCTGGAGCATACTGTCTAGAG   TTTATTCTCAAGTGCTGA               GGACCACTGCAGTTTCAAGTATGGATGGTGTTGAGCTT   GGGCTGACCTGGAGCATA               TGGGGGATGCTAGATGCATCTATAAATGTGTGCTGTTT   CTGTCTAGAGGGACCACT               ACTTTTTTTCATCTTGGTATATAATCTTTATTTGTACT   GCAGTCTTCAAGTATGGA               TTGTCAGTTTCATCTGTGTAGAAGATGCCGCGCCTGCA   TGGTGTTGAGCTTTGGGG               GCAGAAATGGTTGAACTCCCGAGAGTGTCCTACACTTA   GTGCTAGATGCATCTATA               GGGGAGAAGCAGCCAAGGGGTTGTTTCCCACCAAGGAC   AATGTGTGCTGTTTACTT               GACCCGTCTGCGCACAAACGGATGAGCCCATCAAGGGC   TTTTTCATCTTGGTATAT               GATTC   AATCTTTATTTGTACTTT                   GTCAGTTTCATCTGTGTA                   GAAGAT{circumflex over ( )}               IST000034   MM0018C   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATGTT   CATGTTCCTTTGTCAAGG   100   unknown               CCTTTGTCAAGGTCTATGGACATTTCTGTCTTTTGGAA   TCTATGGACATTTCTGTC               TGTTCCTCCTTTAGATATCCTGGGGTTTTGTTGTTGTT   TTTTGGAATGTTCCTCCT               GTTTTGTTTTGAGCAGTTGCCGCGCCTGCAGCAGAAAT   TTAGATATCCTGGGGTTT               GGTTGAACTCCCGAGAGTGTCCTACACTTAGGGGAGAA   TGTTGTTGTTGTTTTGTT               GCAGCCAAGGGGTTGTTTCCCACCAAGGACGACCCGTC   TTGAGGCAGT{circumflex over ( )}               TGCGCACAAACGGATGAGCCCATCAAGGCGAATTC               IST000035   MM0019B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATGAG   CATGAGCTGTCGATAGTG   129   unknown               CTGTCGATAGTGACCTGCAGTCAAGGAAATCTGAGGGC   ACCTGCAGTCAAGGAAAT               TTCCTAATTAACAGAGGAGCTCTAAATGAGAGTAACGC   CTGAGGGCTTCCTAATTA               GCTCCACAAACCCCTCACACTCGGTAAGTGTCACGGTG   ACAGAGGAGCTCTAAATG               CAGATAATTGCCGCGCCTGCAGCAGAAATGGTTGAACT   AGAGTAACGCGCTCCACA               CCCGAGAGTGTCCTACACTTAGGGGAGAAGCAGCCAAG   AACCCCCTCACACTCGGT               GGGTTGTTTCTCACCAAGGACGACCCTCTGCGCACAAA   AAGTGTCACGGTGCAGAT               CGGATGAGCCCATCAAGGGCGAATTC   AT{circumflex over ( )}               IST000036   MM0021B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATGAG   CATGAGCACTAAAGGCAG   105   unknown               CACTAAAGGCAGCTAACCAAAACAGAATCCCACAGACT   CTAACCAAAACAGAATCC               ACTCTGCAGCTTCTTAGGCTGGCTCGTTTTTCCAGTCT   CACAGACTACTCTGCAGC               AGCTATTGCCTTATTCCTCTTCTGCCGCGCCTACAACA   TTCTTAGGCTGGCTCGTT               GAAATGGTTGAACTCCCGAGAGTGTCCTACACTTAGGG   TTTCCAGTCTAGCTATTG               GAGAAGCAGCCAAGGGGTTGTTTCCCACCAAGGACGAC   CCTTATTTCCTCTTC{circumflex over ( )}               CCGTCTGCGCACAAACGGATGAGCCCTCAAGGGCGAAT               TC               IST000037   MM0021C   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATGGA   CATGGATAGAGGATGGAA   66   unknown               TAGAGGATGGAAGTTGAAAACCTGCTATTAAGAATATA   GTTGAAAACCTGCTATTA               GCCCTGTCCATTAGTGAGAGTGTGCCGCGCCTGCAGCA   AGAATATAGCCCTGTCCA               GAAATGGTTGAACCCCGAGAGTGTCCTACACTTAGGGG   TTAGTGAGAGTG{circumflex over ( )}               AGAAGCAGCCAAGGGGTTGTTTCCCACCAAGGACGACC               CGTCTGCGCACAAACGGATGAGCCCATCAAGGGCGAAT               TC               IST000038   MM0024B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATGAGGG   CATGAGGGGTACAAATT   78   unknown               GTACAAATTTGTTGTGCCTAAAAGCATTGAAGTGTAGGGA   TGTTGTGCCTAAAAGCA               AGACTGGGAAGTGGCACGGTCAGACTTAGGTGCCGCGCCT   TTGAAGTGTAGGGAAGA               GCAGCAGAATGGTTGAACTCCCGAGAGTGTCCTACACTTA   CTGGGAAGTGGCACGGT               GGGGAGAAACAGCCAAGGGGTTGTTTCCCACCAAGGACGA   CAGACTTAGG{circumflex over ( )}               CCCGTCTGCGCACAAACGGATGAGCCCATCAAGGGCGAAT               TC               IST000039   MM0024C   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATGTCAC   CATGTCACACTGGGGAC   46   unknown               ACTGGGGACTCCCTCCGGGGAGGGAGCAGGCTGGGGGGTG   TCCCTCCGGGGAGGGAG               CCGCGCCTGCAGCAGAAATGGTTGAACTCCCGAGAGTGTC   CAGGCTGGGGGG{circumflex over ( )}               CTACACTAGGGGAGAAGCAGCCGAGGGGTTGTTTCCCACC               AAGGACGACCCGTCTGCGCACAAACGGATGAGCCCATCAA               GGGCGAATTC               IST000040   MM0022B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATGGCTA   CATGGCTATCCTTTCCT   99   Wnt1               TCCTTTCCTTTCATTGCTGCTACCTCCCCAGCTCTTGCAC   TTCATTGCTGCTACCTC               TAGGAGAGCTGGACTGTTGGTGTGCTGAGTCCCCATCAGG   CCCAGCTCTTGCACTAG               TGACACATTGTGCCGCGCCTGCAGCAGAAATGGTTGAACT   GAGAGCTGGACTGTTGG               CCCGAGAGTGTCCTACACTTAGGGGAGAAGCAGCCAAGGG   TGTGCTGAGTCCCCATC               GTTGTTTCCCACCAAGGACGACCCGTCTGCGCACAAACGG   AGGTGACACACTTG{circumflex over ( )}               ATGAGCCCATCAAGGCGAATTC               IST000041   MM0022C   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATGTCAG   CATGTCAGGTGGCTCAC   85   unknown               GTGGCTCACCTGCAGAGGATCTAATTAATGCCTCCACCCT   CTGCAGAGGATCTAATT               CCAGGAGTGCCTACCCTCACATAAACATATCCATATATGC   AATGCCTCCACCCTCCA               CGCGCCTCAGCAGAAATGGTTGAACTCCCGAGAGTGTCCT   GGAGTGCCTACCCTCAC               ACACTTAGGGGAGAAGCAGCCAAGGGGTTGTTTCCCACCA   ATAAACATATCCATATA{circumflex over ( )}               AGGACGACCCGTCTGCGCACAAACGGATGAGCCCATCAAG               GGCGAATTC               IST000042   MM0022D   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATGTCTG   CATGTCTGCTGTCTAAA   72   unknown               CTGTCTAAACCTCCTAATCTATATATTTTGTTACAGATGT   CCTCCTAATCTATATAT               TCAAGATGACTACAGTCCACATATTGCCGCGCCTGCAGCA   TTTGTTACAGATGTTCA               GAAATGGTGAACTCCCGAGAGTGTCCTACACTTAGGGGAG   AGATGACTACAGTCCAC               AAGCAGCCAAGGGGTTGTTTCCCACCAAGGACGACCCGTC   ATAT{circumflex over ( )}               TGCGCACAAACGGATGAGCCCATCAAGGGCGAATTC               IST000043   MM0022F   GAATTCGCCCTTCGTGGTACACTATCAGTAGGCATGGTTC   CATGGTTCTGGGAA{circumflex over ( )}   14   too short               TGGGAATGCCGCGCCTGCAGCAGAAATGGTTGAACTCCCG               AGAGTGTCCTACACTTAGGGGAGAAGCAGCCAAGGGGTTG               TTTCCCACAAGGACGACCCGTCTGCGCACAAACGGATGAG               CCCATCAAGGGCGAATTC               IST000044   MM0022G   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATGTGCC   CATG{circumflex over ( )}   4   too short               GCGCCTGCAGCAGAAATGGTTGAACTCCCGAGAGTGTCCT               ACACTTAGGGGAGAAGCAGCCAAGGGGTTGTTTCCCACCA               AGGACGACCGTCTGCGCACAAACGGATGAGCCCATCAAGG               GCGAATTC               IST000045   MM0017B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATGTGTC   CATGTGTCACTCAAAGG   205   unknown               ACTCAAAGGCTGCTGAGAATCAGGCTGTACCTGCATTCCT   CTGCTGAGAATCAGGCT               AAGCCATCCACAGCCATCCTGACCCACAGCAAATGCTGGC   GTACCTGCATTCCTAAG               AGTCGCCCACAGCTGGACTCCGTTCCTCCCTCCACTCCTA   CCATCCACAGCCATCCT               TAGCCGAGGCTATCCACACAGGCTATTTCAGTGCCCTAAG   GACCCACAGCAAATGCT               CCTTGCTACCCTTATGTATACATTGAGGACAATGACTGCC   GGCAGTCGCCCCACAGC               GCGCCTGCAGCAGAATGGTTGAACTCCCGAGAGTGTCCTA   TGGACTCCGTTCCTCCC               CACTAAGGGGAGAAGCAGCCAAGGGGTTGTTTCCCACCAA   TCCACTCCATAGCCGAG               GGACGACCCGTCTGCGCACAAACGGATGAGCCCATCAAGG   GCTATCCACACAGGCTA               GCGAATTC   TTTCAGTGCCCTAAGCC                   TTGCTACCCTTATGTAT                   ACATTGAGGACAATGAC{circumflex over ( )}               IST000046   MM0017D   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATGGAA   CATGGAAATA{circumflex over ( )}   10   too short               ATATGCCCCGCCTGCAGCAGAAATGGTTGAACTCCCGAG               AGTGTCCTACACTTAGGGGAGAAGCAGCCAAGGGGTTGT               TTCCCACCAAGACGACCCGTCTGCGCACAAACGGATGAG               CCCATCAAGGGCGAA~TTC               IST000047   MM0017E   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATGCCG   CA{circumflex over ( )}   2   too short               CGCCTGCAGCAGAAATGGTTGAACTCCCGAGAGTGTCCT               ACACTTAGGGGAGAAGCAGCCAAGGGGTTGTTTCCCACC               AAGGACGACCGTCTGCGCACAAACGGATGAGCCCATCAA               GGGCGAATTC               IST000048   MM0020B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATGGAG   CATGGAGAAACCTGGGCT   126   unknown               AAACCTGGGCTTATTCAAGCAGTTTCCTTTGTTTACCCT   TATTCAAGCAGTTTCCTT               GCCCAGGGTTGCCAGTGAAGGGGCTCCTCCATCACTAAC   TGTTTACCCTGCCCAGGG               TAAAGGTCTTTCCTATGCTGGTTCCTCTCCACCCCACCA   TTGCCAGTGAAGGGGCTC               TTGCCGCGCCTGCAGCAGAAATGGTTGAACTCCCGAGAG   CTCCATCACTAACTAAAG               TGTCCTACACTTAGGGGAGAAGCAGCCAAGGGGTTGTTT   GTCTTATCCTATGCTGGT               CCCACCAAGGACGACCCGTCGCGCACAAACGGATGAGCC   TCCTCTCCACCCCACCAT{circumflex over ( )}               CATCAAGGGCGAATTC               IST000049   MM0020D   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATGGGT   CATGGGTGTG{circumflex over ( )}   10   too short               GTGTGCCGCGCCTGCAGCAGAAATGGCTGAACTCCCGAG               AGTGTCCTACACTTAGGGGAGAAGCAGCCAAGGGGTTGT               TTCCCACCAAGACGACCCGTCTGCGCACAAACGGATGAG               CCCATCAAGGGCGAATTC               IST000050   MM0023B   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATGTCC   CATGTCCTCCACAGCCGA   221   Fgf3               TCCACAGCCGAGCACCCTCAACTGTCTCCCAGGGCCTCT   GCACCCTCAACTGTCTCC               GTTCTAGCCAGGGTCTGCAGGGTCTCTGCCCCACGCCTA   CAGGGCCTCTGTTCTAGC               GCCCCTGAGAATCTTAAGCGGTCTGAAAACTACGCCACT   CAGGGTCTGCAGGGTCTC               GAACTGCTAAAACCCTGGAGTCACTGATGGAACCTGCTT   TGCCCCACGCCTAGCCCC               TACAGGGGACAGCCCTGTGTCTCGAAGACCAACCTAAGC   TGAGAAATCTTAAGCGGT               TCCTAAGATGTCACAAATTGCGCGCCTGCAGCAGAAATG   CTGAAAACTACGCCACTG               GTTGAACTCCCGAGAGTGTCCTACACTTAGGGGAGAAGC   ACTGCTAAAACCCTGGAG               AGCCAAGGGGTTGTTTCCCACCAAGGACGACCCGTCTGC   TCACTGATGGAACCTGCT               GCACAAACGGATGAGCCCATCAAGGGCGAATC   TTACAGGGGACAGCCCTG                   TGTCTCGAAGACCAACCT                   AAGGTCCTAAGATGTCAC                   AAAT{circumflex over ( )}               IST000051   MM0023C   GAATTCGCCCTTCGTGCTACACTATCAGTAGGCATGACG   CATGACGACTTGAAAAAT   39   unknown               ACTTGAAAAATGACGAAATCACTAAAAAACAATGCCGCG   GACGAAATCACTAAAAAA               CCTGCAGCAGAAATGGTTGAACTCCCGAGAGTGTCCTAC   CAA{circumflex over ( )}               ACTTAGGGGAAAGCAGCCAAGGGGTTGTTTCCCACCAAG               GACGACCCGTCTGCGCACAAACGGATGAGCCCATCAAGG               GCGAATTC                  
 
         [0062]     It is understood that the examples and embodiments described herein are for illustrative purposes only and that various modifications or changes in light thereof will be suggested to persons skilled in the art and are to be included within the spirit and purview of this application and scope of the appended claims. All publications, patents, and patent applications cited herein are hereby incorporated by reference in their entirety for all purposes.